Antiviral phosphodiamide prodrugs of tenofovir

ABSTRACT

Compounds of Formula I and pharmaceutically acceptable salts and co-crystals thereof are useful for the inhibition of HIV reverse transcriptase. The compounds may also be useful for the prophylaxis or treatment of infection by HIV and in the prophylaxis, delay in the onset or progression, and treatment of AIDS. The compounds and their salts can be employed as ingredients in pharmaceutical compositions, optionally in combination with other antiviral agents, immunomodulators, antibiotics or vaccines.

RELATED APPLICATIONS

This application claims priority to EP Application No. EP15306977.8filed Dec. 10, 2015 and to PCT/US2016/064883 filed Dec. 5, 2016. Each ofthe aforementioned applications to which this application claimspriority is herein incorporated by reference in its entirety.

BACKGROUND OF THE INVENTION

The retrovirus designated human immunodeficiency virus (HIV),particularly the strains known as HIV type-1 (HIV-1) and type-2 (HIV-2),have been etiologically linked to the immunosuppressive disease known asacquired immunodeficiency syndrome (AIDS). HIV seropositive individualsare initially asymptomatic but typically develop AIDS related complex(ARC) followed by AIDS. Affected individuals exhibit severeimmunosuppression which makes them highly susceptible to debilitatingand ultimately fatal opportunistic infections. Replication of HIV by ahost cell requires integration of the viral genome into the host cell'sDNA. Since HIV is a retrovirus, the HIV replication cycle requirestranscription of the viral RNA genome into DNA via an enzyme known asreverse transcriptase (RT).

Reverse transcriptase has three known enzymatic functions: The enzymeacts as an RNA-dependent DNA polymerase, as a ribonuclease, and as aDNA-dependent DNA polymerase. In its role as an RNA-dependent DNApolymerase, RT transcribes a single-stranded DNA copy of the viral RNA.As a ribonuclease, RT destroys the original viral RNA and frees the DNAjust produced from the original RNA. During the viral RNA-dependentpolymerization process, RT's ribonuclease activity is required forremoving RNA and leaving the polypurine tract preserved for initiationof DNA-dependent polymerization. As a DNA-dependent DNA polymerase, RTmakes a second, complementary DNA strand using the first DNA strand as atemplate. The two strands form double-stranded DNA, which is integratedinto the host cell's genome by HIV integrase.

It is known that compounds that inhibit enzymatic functions of HIV RTwill inhibit HIV replication in infected cells. These compounds areuseful in the treatment of HIV infection in humans. There are twoclasses of RT inhibitors: one is non-nucleoside active site competitiveRT inhibitors (NNRTIs), such as efavirenz (EFV), nevirapine (NVP),etravirine (ETR), and rilpivirine (RPV), and the other is active site RTinhibitors which include nucleoside reverse transcriptase inhibitors(NsRTIs) and nucleotide reverse transcriptase inhibitors (NtRTIs),collectively referred to as NRTIs. Examples of NsRTI's include3′-azido-3′-deoxythymidine (AZT), 2′,3′-dideoxyinosine (ddI),2′,3′-dideoxycytidine (ddC), 2′,3′-didehydro-2′,3′-dideoxythymidine(d4T), 2′,3′-dideoxy-3′-thiacytidine (3TC), abacavir, and emtricitabine.Examples of NtRTIs include tenofovir (TFV, also known as PMPA,9-(2-phosphonyi-methoxypropyl)adenine), tenofovir disoproxil fumarate(VIREAD®, U.S. Pat. Nos. 5,977,089, 5,935,946) and tenofovir alafenamidefumarate (U.S. Pat. Nos. 7,390,791, 8,754,065).

TFV belongs to a class of HIV anti-retroviral (ARV) agents known asnucleotide analog reverse transcriptase inhibitors (NRTIs). Tenofovir isa monophosphonate:

After ebing taken up by cells, TFV is first converted totenofovir-monophosphate (TFV-MP) by adenosine monophosphate kinase andthen to the active antiviral tenofovir-diphosphate (TFV-DP) by5′-nucleoside diphosphate kinase.

TFV-DP inhibits HIV DNA synthesis by competing with the naturalsubstrate, deoxyadenosine triphosphate, for incorporation into thecomplementary DNA strand by HIV to reverse transcriptase; followingincorporation, TFV acts as a chain terminator due to lack of a3′-hydroxyl group that is required for addition of the next nucleotide.TFV has poor cellular permeability and thus has limited bioavailability.Tenofovir disoproxil fumarate (TDF) is approved for treating HIVinfection and is marketed by Gilead under the trade name VIREAD™. Thedisoproxil prodrug improves cell permeability and absorption after oraldosing, with the pro-moiety being cleaved rapidly after absorption toyield the parent TFV. As a result, the circulating level of TFV is muchhigher than that of TDF. Tenofovir alafenamide fumarate (TAF) iscurrently approved by the USFDA as an active ingredient in combinationwith additional ARVs for treating HIV infection in the pharmaceuticalproducts GENVOYA®, ODEFSEY® and DESCOVY®.

While each of the foregoing drugs is effective in treating HIV infectionand AIDS, there remains a need to develop additional HIV antiviral drugsincluding additional RT inhibitors. A particular problem is thedevelopment of mutant HIV strains that are resistant to the knowninhibitors. The use of RT inhibitors to treat AIDS often leads toviruses that are less sensitive to the inhibitors. This resistance istypically the result of mutations that occur in the reversetranscriptase segment of the pol gene. The continued use of antiviralcompounds to prevent HIV infection will inevitably result in theemergence of new resistant strains of HIV. Accordingly, there is aparticular need for new RT inhibitors that are effective against mutantHIV strains.

SUMMARY OF THE INVENTION

The present invention is directed to phosphodiamide prodrugs oftenofovir and their use in the inhibition of nucleotide reversetranscriptase. In addition to the use of said compounds in theinhibition of HIV reverse transcriptase, the invention is also directedto the use of said compounds for prophylaxis of infection by HIV, thetreatment of infection by HIV, and the prophylaxis, treatment, and/ordelay in the onset or progression of AIDS and/or ARC. The invention alsoencompasses methods of using said compounds for the treatment of chronichepatitis B infection.

DETAILED DESCRIPTION OF TI-IE INVENTION

The present invention is directed to compounds of structural Formula I:

or pharmaceutically acceptable salts thereof, wherein:

-   R^(A) is a D-amino acid ester residue of formula (i) or a D-proline    ester residue of formula (ii):

-   R¹ is (a) H, or (b) —C₁₋₃alkyl optionally substituted with one to    three substituents independently selected from fluoro, chloro,    bromo, and —OH;-   R² is (a) H, (b) —C₁₋₄alkyl, (c) —C₁₋₄alkyl substituted with —OH,    —SH, —SCH₃, —NH₂ or —NH—C(═NH)—NH₂, (d) —CH₂-phenyl, (e)    —CH₂-phenol, (f) —(CH₂)₁₋₂—COOH, (g) —(CH₂)₁₋₂—CONH₂, (h)    —CH₂-1H-indole, (i) —CH₂-imidazole, (j) aryl (for example but not    limited to phenyl or naphthyl) or (k) heteroaryl (for example but    not limited to pyridine);-   R³ is-   (a) —C₁₋₁₀alkyl unsubstituted or substituted with one to three    substituents independently selected from fluoro, chloro, bromo,    —OR⁷, —SH, —NR⁸R⁹, —C₃₋₆cycloalkyl or spiro—C₃₋₆cycloalkyl,-   (b) —CH₂-phenyl unsubstituted or substituted with one to three    substituents independently selected from fluoro, chloro, bromo,    —OR¹⁰, —SH, —NR⁸R⁹ or —C₁₋₃alkyl,-   (c) —C₃₋₈cycloalkyl unsubstituted or substituted with one to three    substituents independently selected from fluoro, chloro, bromo,    —OR¹⁰, —SH, —NR⁸R⁹ or —C₁₋₃alkyl,-   (d) aryl unsubstituted or substituted with one to three substituents    independently selected from fluoro, chloro, bromo, —OR¹⁰, —SH,    —NR⁸R⁹ or —C₁₋₃alkyl,-   (e) —C₁₋₅alkyl-X—C₁₋₅alkyl wherein X is O, S, or NH,-   (f) heteroaryl unsubstituted or substituted with one to three    substituents independently selected from fluoro, chloro, bromo,    —OR¹⁰, —SH, —NR⁸R⁹ or —C₁₋₃alkyl, or-   (g) a heterocyclic ring unsubstituted or substituted with one to    three substituents independently selected from fluoro, chloro,    bromo, —OR¹⁰, —SH, —NR⁸R⁹ or —C₁₋₃alkyl;-   R^(B) is a geminally di-substituted amino acid ester residue of    formula (iii):

-   R⁴ and R⁵ are each independently selected from (a) —C₁₋₄alkyl, (b)    —C₁₋₄alkyl substituted with —OH, —SH, —SCH₃, —NH₂,    —NH—C(═NH)—NH₂, (c) —CH₂-phenyl, (d) —CH₂phenol, (e)    —(CH₂)₁₋₂—COOH, (f) —(CH₂)₁₋₂CONH₂, (g) —CH₂-1H-indole, (h)    —CH₂-imidazole, (i) aryl (for example but not limited to phenyl or    naphthyl) or (j) heteroaryl (for example but not limited to    pyridine); or R⁴ and R⁵ are joined together with the carbon to which    they are both attached to form —C₃₋₆cycloalkyl or a 4 to 6-membered    heterocyclic ring;-   R⁶ is-   (a) —C₁₋₁₀alkyl unsubstituted or substituted with one to three    substituents independently selected from fluoro, chloro, bromo,    —OR¹¹, —SH, —NR¹²R¹³, —C₃₋₆cycloalkyl or spiro-C₃₋₆cycloalkyl,-   (b) —CH₂-phenyl unsubstituted or substituted with one to three    substituents independently selected from fluoro, chloro, bromo,    —OR¹⁴, —SH, —NR¹²R¹³ or —C₁₋₃alkyl,-   (c) —C₃₋₈cycloalkyl unsubstituted or substituted with one to three    substituents independently selected from fluoro, chloro, bromo, —OR    , —SH, —NR¹²R¹³ or —C₁₋₃alkyl,-   (d) aryl unsubstituted or substituted with one to three substituents    independently selected from fluoro, chloro, bromo, —OR , —SH,    —NR¹²R¹³ or —C₁₋₃alkyl,-   (e) —C₁₋₅alkyl-X—C₁₋₅alkyl wherein X is O, S or NH;-   (f) heteroaryl unsubstituted or substituted with one to three    substituents independently selected from fluoro, chloro, bromo,    —OR¹⁴, —SH, —NR¹²R¹³ or —C₁₋₃alkyl, or-   (g) a heterocyclic ring unsubstituted or substituted with one to    three substituents independently selected from fluoro, chloro,    bromo, —OR¹⁴, —SH, —NR¹²R¹³ or —C₁₋₃alkyl;-   R⁷ and R¹¹ are each independently selected from —H or    —C₃₋₆cycloalkyl;-   R⁸ _(and) R⁹ are each independently selected from -—H, —C₁₋₃alkyl or    —C₃₋₆cycloalkyl;-   R¹² and R¹³ are each independently selected from —H, —C₁₋₃alkyl or    —C₃₋₆cycloalkyl; and-   R¹⁰ and R¹⁴ are each independently selected from —H, —C₁₋₃alkyl or    —C₃₋₆cycloalkyl.

In an embodiment of this invention are compounds of Formula I or thepharmaceutically acceptable salts thereof, wherein R^(A) is formula (i):

referred to herein as compounds of Formula I-i;and the remaining variables are as defined in Formula I.

In an embodiment of this invention are compounds of Formula I or thepharmaceutically acceptable salts thereof, wherein R^(A) is formula(ii):

referred to herein as compounds of Formula I-ii;and the remaining variables are as defined in Formula I.

In Embodiment 1 of this invention are compounds of Formula I, I-i, orI-ii, or Embodiment 2, 2a, 3, 4, 4a, 5, 6, 7, 8, 9 or 10 or a classthereof, or the pharmaceutically acceptable salts thereof, wherein ifpresent (as in Formula I-i) R¹ is H or —C₁₋₄alkyl and R² is H or—C₁₋₄alkyl; and the remaining variables are as defined in Formula I. Ina class thereof, one of R¹ and R² is H, and the other is H or—C₁₋₄alkyl; and in a sub-class thereof one of R¹ and R² is H, and theother is —C₁₋₄alkyl, for example methyl or i-propyl. In a class thereof,R¹ is H or —C₁₋₄alkyl and R² is —C₁₋₄alkyl. In a sub-class thereof, R¹is H and R² is methyl. In a sub-class thereof, R¹ is methyl and R² ismethyl.

In Embodiment 2 of this invention are compounds of Formula I, or I-ii orEmbodiment 1, or the pharmaceutically acceptable salts thereof, whereinR³ is

-   (a) —C₁₋₈alkyl, —CH₂CH₂OH, —CH₂CH₂CH₂OH, —CH₂CH₂SH, —CH₂CH₂CH₂SH,    —CH₂CH₂NH₂, or —CH₂CH₂CH₂NH₂,-   (b) —CH₂-phenyl, unsubstituted or substituted with one to three    substituents independently selected from fluoro, chloro, bromo,    —OR¹⁰, —SH, —NR⁸R⁹ or —C₁₋₃alkyl,-   (c) —C₃₋₆cycloalkvl, unsubstituted or substituted with one to three    substituents independently selected from fluor, chloro, bromo,    —OR¹⁰, —SH, —NR⁸R⁹ or —C₁₋₃alkyl,-   (d) phenyl or naphthyl, each unsubstituted or substituted with one    to three substituents independently selected from fluoro, chloro,    bromo, —OR¹⁰, —SH, —NR⁸R⁹ or —C₁₋₃alkyl,-   (e) —CH₂CH₂OCH₃, —CH₂CH₂CH₂OCH₃, —CH₂CH₂SCH₃, —CH₂CH₂CH₂SCH₃,    —CH₂CH₂NHCH₃, —CH₂CH₂CH₂NHCH₃,-   (f) pyridyl, unsubstituted or substituted with one to three    substituents independently selected from fluoro, chloro, bromo,    —OR¹⁰, —SH, —NR⁸R⁹ or —C₁₋₃alkyl, or-   (g) piperidinyl, pyrrolidinyl, tetrahydrofuranyl, or    tetrahydropyranyl, each unsubstituted or substituted with one to    three substituents independently selected from fluoro. chloro,    bromo, —OR¹⁰, —SH, —NR⁸R⁹ or —C₁₋₃alkyl;    and the remaining variables are as defined in Formula I.    In a class of Embodiment 2, referred to as Embodiment 2a, R³ is    cyclopropyl, cyclobutyl, cyclopentyl or cyclohexyl, and in a    sub-class thereof, R³ is —C₃₋₈alkyl, for example i-propyl; and the    remaining variables are as defined in Formula I.

In Embodiment 3 of this invention are compounds of Formula I, I-i orI-ii or Embodiment 1, 2 or 2a, or the pharmaceutically acceptable saltsthereof, wherein R⁴ and R⁵ are each independently selected from —CH₃,—CH₂CH₃, —C₃alkyl or —C₄alkyl; and the remaining variables are asdefined in Formula I. In another class of Embodiment 3, R⁴ and R⁵ areboth the same atom moiety which is selected from —CH₃, —CH₂CH₃, —C₃alkylor —C₄alkyl; and in a sub-class thereof R⁴ and R⁵ are both —CH₃.

In Embodiment 4 of this invention are compounds of Formula I, I-i orI-ii or Embodiment 1, 2, 2a or 3, or the pharmaceutically acceptablesalts thereof, wherein R⁶ is

-   (a) —C₁₋₈alkyl, —CH₂CH₂OH, —CH₂CH₂CH₂OH, —CH₂CH₂SH, —CH₂CH₂CH₂SH,    —CH₂CH₂NH₂, —CH₂CH₂CH₂NH₂.-   (b) —CH₂-phenyl, unsubstituted or substituted with one to three    substituents independently selected from fluoro, chloro, bromo    —OR¹⁴, —SH, —NR¹²R¹³ or —C₁₋₃alkyl,-   (c) —C₃₋₆cycloalkyl, unsubstituted or substituted with one to three    substituents independently selected from fluoro, chloro, bromo,    —OR¹⁴, —SH, —NR¹²R¹³ or —C₁₋₃alkyl,-   (d) phenyl or naphthyl, each unsubstituted or substituted with one    to three substituents independently selected from fluoro, chloro,    bromo, —OR¹⁴, —SH, —NR¹²R¹³ or —C₁₋₃alkyl,-   (e) —CH₂CH₂OCH₃, —CH₂CH₂CH₂OCH₃, —CH₂CH₂SCH₃, —CH₂CH₂CH₂SCH₃,    —CH₂CH₂NHCH₃, —CH₂CH₂CH₂NHCH₃,-   (f) pyridyl, unsubstituted or substituted with one to three    substituents independently selected from fluoro, chloro, bromo,    —OR¹⁴, —SH, —NR12R¹³ or —C₁₋₃alkyl, or-   (g) piperidinyl, pyrrolidinyl, tetrahydrofuranyl, or    tetrahydropyranyl, each unsubstituted or substituted with one to    three substituents independently selected from fluoro, chloro,    bromo, —OR¹⁴, —SH, —NR¹²R¹³ or —C₁₋₃alkyl;    and the remaining variables are as defined in Formula I.    In a class of Embodiment 4, referred to as Embodiment 4a, R⁶ is    —C₁₋₈alkyl, cyclopropyl, cyclobutyl, cyclopentyl or cyclohexyl, and    in a sub-class thereof R⁶ is —C₃₋₈alkyl, cyclobutyl, cyclopentyl or    cyclohexyl; and the remaining variables are as defined in Formula I.

In Embodiment 5 of this invention are compounds of Formula I, or apharmaceutically acceptable salt thereof, wherein:

-   R¹, is present (as in Formula I-i), is H or —C₁₋₄alkyl;-   R², if present (as in Formula I-i), is H or —C₁₋₄alkyl;-   R³ is-   (a) —C₁₋₈alkyl, —CH₂CH₂OH, —CH₂CH₂CH₂OH, —CH₂CH₂SH, —CH₂CH₂CH₂SH,    —CH₂CH₂NH₂, —CH₂CH₂CH₂NH₂,-   (b) —CH₂-phenyl, unsubstituted or substituted with one to three    substituents independently selected from fluoro, chloro, bromo,    —OR¹⁰, —SH, —NR⁸R⁹ or —C₁₋₃alkyl,-   (c) —C₃₋₆cycloalkyl, unsubstituted or substituted with one to three    substituents independently selected from fluoro, chloro, bromo,    —OR¹⁰, —SH, —NR⁸R⁹ or —C₁₋₃alkyl,-   (d) phenyl or naphthyl, each unsubstituted or substituted with one    to three substituents independently selected from fluoro, chloro,    bromo, —OR¹⁰, —SH, —NR⁸R⁹ or —C₁₋₃alkyl,-   (e) —CH₂CH₂OCH₃, CH₂CH₂CH₂OCH₃, —CH₂CH₂SCH₃, —CH₂CH₂CH₂SCH₃,    —CH₂CH₂NHCH₃, —CH₂CH₂CH₂NHCH₃,-   (f) pyridyl, unsubstituted or substituted with one to three    substituents independently selected from fluoro, chloro, bromo,    —OR¹⁰, —SH, —NR⁸R⁹ or —C₁₋₃alkyl, or-   (g) piperidinyl, pyrrolidinyl, tetrahydrofuranyl, or    tetrahydropyranyl, each unsubstituted or substituted with one to    three substituents independently selected from fluoro, chloro,    bromo, —OR¹⁰, —SH, —NR⁸R⁹ or —C₁₋₃alkyl, or-   R⁴ and R⁵ are both the same moiety which is selected from —CH₃,    —CH₂CH₃, —C₃alkyl or —C₄alkyl;-   R⁶ is-   (a) —CH₁₋₈alkyl, —CH₂CH₂OH, —CH₂CH₂CH₂OH, —CH₂CH₂SH, —CH₂CH₂CH₂SH,    —CH₂CH₂NH₂, —CH₂CH₂CH₂NH₂,-   (b) —CH₂-phenyl, unsubstituted or substituted with one to three    substituents independently selected from fluoro, chloro, bromo,    —OR¹⁴, —SH, —NR¹²R¹³ or —C₁₋₃alkyl,-   (c) —C₃₋₆cycloalkyl, unsubstituted or substituted with one to three    substituents independently selected from fluoro, chloro, bromo,    —OR¹⁴, —SH, —NR¹²R¹³ or —C₁₋₃alkyl,-   (d) phenyl or naphthyl, each unsubstituted or substituted with one    to three substituents independently selected from fluoro, chloro,    bromo, —OR¹⁴, —SH, —NR¹²R¹³ or —C₁₋₃alkyl,-   (e) —CH₂CH₂OCH₃, —CH₂CH₂CH₂OCH₃, —CH₂CH₂SCH₃, —CH₂CH₂CH₂SCH₃,    —CH₂CH₂NHCH₃, —CH₂CH₂CH₂NHCH₃,-   (f) pyridyl, unsubstituted or substituted with one to three    substituents independently selected from fluoro, chloro, bromo,    —OR¹⁴, —SH, —NR¹²R¹³ or —C₁₋₃alkyl,-   (g) piperidinyl, pyrrolidinyl, tetrahydrofuranyl, or    tetrahydropyranyl, each unsubstituted or substituted with one to    three substituents independently selected from fluoro, chloro,    bromo, —OR¹⁴, —SH, —NR¹²R¹³ or —C₁₋₃alkyl;    and the remaining variables are as defined in Formula I.

In Embodiment 6 of this invention are compounds of Formula I, I-i orI-ii, or a pharmaceutically acceptable salt thereof, wherein

-   R¹, if present (as in Formula I-i), is H or —C₁₋₄alkyl;-   R², if present (as in Formula I-i), is —C₁₋₄alkyl; and-   R³ is —C₁₋₈alkyl, cyclopropyl, cyclobutyl, cyclopentyl or    cyclohexyl, or in a class of-   Embodiment 6, R³ is i-propyl;    and the remaining variables are as defined in Formula I.

In Embodiment 7 of this invention are compounds of Embodiment 6 havingFormula I or I-i, or a pharmaceutically acceptable salt thereof, wherein

-   R⁶ is —C₁₋₈alkyl, cyclopropyl, cyclobutyl, cyclopentyl or    cyclohexyl, or in a class of Embodiment 7, R⁶ is —C₃₋₈alkyl;    and the remaining variables are as defined in Formula I.

In Embodiment 8 of this invention are compounds of Embodiment 6 havingFormula I or I-ii, or a pharmaceutically acceptable salt thereof,wherein

-   R⁶ is —C₁₋₈alkyl, cyclopropyl, cyclobutyl, cyclopentyl or    cyclohexyl, or in a class of Embodiment 8, R⁶ is —C₃₋₈alkyl;    and the remaining variables are as defined in Formula I.

In Embodiment 9 of this invention are compounds of Formula I, or apharmaceutically acceptable salt thereof, wherein:

-   R¹, if present (as in Formula I-i), is H or —CH₃;-   R², if present (as in Formula I-i), is —C₁₋₄alkyl;-   R³ is —C₁₋₈alkyl, cyclopropyl, cyclobutyl, cyclopentyl or    cyclohexyl;-   R⁴ and R⁵ are both —CH₃;-   R⁶ is —C₁₋₈alkyl, cyclopropyl, cyclobutyl, cyclopentyl or    cyclohexyl;    and the remaining variables are as defined in Formula I.

In Embodiment 10 of this invention are compounds of Formula I, or apharmaceutically acceptable salt thereof, wherein:

-   R¹, if present (as in Formula I-i), is H or —CH₃;-   R², if present (as in Formula I-i), is —C₁₋₄alkyl;-   R³ is —C₃₋₈alkyl, cyclobutyl, cyclopentyl or cyclohexyl;-   R⁴ and R⁵ are both —CH₃;-   R⁶ is —C₃₋₈alkyl, cyclobutyl, cyclopentyl or cyclohexyl;    and the remaining variables are as defined in Formula I.

In Embodiment 11 of this invention are compounds of Formula I, or apharmaceutically acceptable salt thereof, wherein:

-   R¹, if present (as in Formula I-i), is H or —CH₃;-   R², if present (as in Formula I-i), is —C₁₋₄alkyl;-   R³ is —C₃₋₈alkyl, or in a class of Embodiment 11, R³ is i-propyl;-   R⁴ and R⁵ are both —CH₃;-   R⁶ is —C₃₋₈alkyl, cyclobutyl, cyclopentyl or cyclohexyl;    and the remaining variables are as defined in Formula I.

In Embodiment 12 of this invention are compounds of Formula I, or apharmaceutically acceptable salt thereof, wherein:

-   R¹, if present (as in Formula I-i), is H or —CH₃;-   R², if present (as in Formula I-i), is —C₁₋₄alkyl;-   R³ is —C₃₋₈alkyl, cyclobutyl, cyclopentyl or cyclohexyl;-   R⁴ and R⁵ are both —CH₃;-   R⁶ is —C₃₋₈alkyl;    and the remaining variables are as defined in Formula I.

Reference to the compounds of Formula I herein encompasses the compoundsof Formula I and all embodiments, classes and sub-classes thereof. Thecompounds of the invention encompass compounds of Formula I and saltsthereof when such salts are possible, including pharmaceuticallyacceptable salts.

As used herein, “alkyl” refers to both branched-and straight-chainsaturated aliphatic hydrocarbon groups having the specified number ofcarbon atoms in a specified range. For example the term “C₁₋₁₀alkyl”means linear or branched chain alkyl groups, including all possibleisomers, having 1, 2, 3, 4, 5, !, 8, 9 or 10 carbon atoms, and includeseach of the decyl, nonyl, octyl, heptyl, hexyl and pentyl isomers aswell as n-, iso-, sec-and tert-butyl (butyl, s-butyl, i-butyl, t-butyl;Bu=butyl, collectively “—C₄alkyl”), n-and iso-propyl (propyl, i-propyl,Pr=propyl, collectively “—C₃alkyl”), ethyl (Et) and methyl (Me),“C₁₋₄alkyl” has 1, 2, 3 or 4 carbon atoms, and includes each of n-,iso-, sec-and tert-butyl, n-and i-propyl, ethyl and methyl.

“Cycloalkyl” refers to a cyclized alkyl ring having the indicated numberof carbon atoms in a specified range. Thus, for example, “C₃₋₈cycloalkyl” encompasses each of cyclopropyl, cyclobutyl, cyclopentyl,cyclohexyl, cyloheptyl and cyclooctyl. “C₃₋₆cycloalkyl” encompasses eachof cyclopropyl, cyclobutyl, cyclopentyl and cyclohexyl. When cycloalkylis a substituent on an alkyl group in a compound of Formula 1, thecycloalkyl substituent can be bonded to any available carbon in thealkyl group. The following are illustrations of —C₃₋₆cycloalkylsubstituents wherein the substituent is cyclopropyl in bold:

“Spiro-C₃₋₆cycloalkyl” refers to a cycloalkyl ring bonded to anon-terminal carbon atom wherein the non-terminal carbon atom is sharedwith the cycloalkyl group. Spiro-C₃₋₆cycloalkyl encompasses each ofspiro-cyclopropyl, spiro-cyclobutyl, spiro-cyclopentyl andspiro-cyclohexyl. The following is an illustration of aspino-C₃₋₆cycloalkyl substituent wherein the substituent isspino-cyclopropyl in bold:

Examples of —C₁₋₅alkyl-X—C₁₋₅alkyl groups include, but are not limitedto, —CH₂CH₂OCH₃, —CH₂CH₂CH₂OCH₃, —CH₂CH₂SCH₃, —CH₂CH₂CH₂SCH₃,—CH₂CH₂NHCH₃ or —CH₂CH₂CH₂NHCH₃.

“Aryl” refers to (i) phenyl, (ii) 9-or 10-membered bicyclic, fusedcarbocylic ring systems in which at least one ring is aromatic, and(iii) 11-to 14-membered tricyclic, fused carbocyclic ring systems inwhich at least one ring is aromatic. Suitable aryls include, forexample, substituted and unsubstituted phenyl and substituted andunsubstituted naphthyl. An aryl of particular interest is unsubstitutedor substituted phenyl.

“Heteroaryl” refers to (i) a 5-or 6-membered heteroaromatic ringcontaining from 1 to 4 heteroatoms independently selected from N, O andS, wherein each N is optionally in the form of an oxide, and (ii) a 9-or10-membered bicyclic fused ring system, wherein the fused ring system of(ii) contains from 1 to 6 heteroatoms independently selected from N, Oand S, wherein each ring in the fused ring system contains zero, one ormore than one heteroatom, at least one ring is aromatic, each N isoptionally in the form of an oxide, and each S in a ring which is notaromatic is optionally S(O) or S(O)₂. Examples of 5—memberedheteroaromatic rings include, but are not limited to, pyrrolyl,pyrazolyl, triazolyl (i.e., 1,2,3-triazolyl or 1,2,4-triazolyl),imidazolyl, tetrazolyl, furanyl, furanonyl (e.g., furan-2(5H)-one),thienyl, thiazolyl, isothiazolyl, oxazolyl, isooxazolyl, oxadiazolyl(i.e., the 1,2,3-, 1,2,4-, 1,2,5-(furazanyl), or 1,3,4-oxadiazolylisomer), oxatriazolyl, and thiadiazolyl. Examples of 6-memberedheteroaromatic rings include, but are not limited to, pyridyl (alsoreferred to as pyridinyl), pyrimidinyl, pyrazinyl, pyridazinyl, andtriazinyl. Examples of 9-and 10-membered heteroaromatic bicyclic fusedring systems include, but are not limited to, benzofuranyl, indolyl,indazolyl, naphthyridinyl, isobenzofuranyl, benzopiperidinyl,benzisoxazolyl, benzoxazolyl, chromenyl, quinolinyl, isoquinolinyl,isoindolyl, benzopiperidinyl, benzofuranyl, imidazo[1,2-a]pyridinyl,benzotriazolyl, indazolyl, indolinyl, and isoindolinyl. A class ofheteroaryls includes unsubstituted or substituted pyridyl or pyritnidyl,and particularly unsubstituted or substituted pyridyl.

The term “heterocyclic ring” refers to (i) a saturated 4-to 7-memberedcyclized ring and (ii) an unsaturated, non-aromatic 4 to 7-memberedcyclized ring comprised of carbon atoms and 1-4 heteratoms independentlyselected from O, N and S. Heterocyclic rings within the scope of thisinvention include, for example but are not limited to, azetidinyl,piperidinyl, morpholinyl, thiomorpholinyl, thiazolidinyl,isothiazolidinyl, oxazolidinyl, isoxazolidinyl, pyrrolidinylimidazolidinyl, piperazinyl, tetrahydrofuranyl, tetrahydrothienyl,pyrazolidinyl, hexahydropyrimidinyl, thiazinanyl, thiazepanyl, azepanyl,diazepanyl, tetrahydropyranyl, tetrahydrothiopyranyl, and dioxanyl.Examples of 4-to 7-membered, unsaturated, non-aromatic heterocyclicrings within the scope of this invention include mono-unsaturatedheterocyclic rings corresponding to the saturated heterocyclic ringslisted in the preceding sentence in which a single bond is replaced witha double bond (e.g., a carbon-carbon single bond is replaced with acarbon-carbon double bond).

In a class of heterocyclic rings are 4 to 6-membered saturatedmonocyclic rings comprised of carbon atoms and 1 or 2 heteroatoms,wherein the heteroatoms are independently selected from N, O and S.Examples of 4 to 6 membered heterocyclic rings include but are notlimited to, azetidinyl, pyrrolidinyl, piperidinyl piperazinyl,morpholinyl, thiomorpholinyl, tetrahydrofuranyl, tetrahydropyranyl andtetrahydrothiopyranyl, and a sub-class thereof is piperidinyl,pyrrolidinyl, tetrahydrofuranyl or tethrahdropyranyl. The following is anon-limiting illustration of R4 and R5 when they are joined together toform a heterocyclic ring:

With respect to substituents on a molecule. “germinally” or “geminal”refers to two substituents, which may be the same or different, on onecarbon.

It is understood that the specific rings and ring systems suitable foruse in the present invention are not limited to those listed in thepreceding paragraphs. These rings and ring systems are merelyrepresentative.

As would be recognized by one of ordinary skill in the art, certaincompounds of the present invention maybe able to exist as tautorners.All tautomeric forms of these compounds, whether isolated individuallyor in mixtures, are within the scope of the present invention. Forexample, in instances where an —OH substituent is permitted on aheteroaromatic ring and keto-enol tautomerism is possible, it isunderstood that the substituent might in fact be present, in whole or inpart, in the oxo (═O) form.

A “stable” compound is a compound which can be prepared and isolated andwhose structure and properties remain or can be caused to remainessentially unchanged for a period of time sufficient to allow use ofthe compound for the purposes described herein (e.g., therapeutic orprophylactic administration to a subject). The compounds of the presentinvention are limited to stable compounds embraced by Formula I and itsembodiments. For example, certain moieties as defined in Formula I maybe unsubstituted or substituted, and the latter is intended to encompasssubstitution patterns (i.e., number and kind of substituents) that arechemically possible for the moiety and that result in a stable compound.

Each compound of Formula I is comprised of a phosphodiamide amino acidester having a defined (R) chiral center in the alkyl-ether linkinggroup which connects the nucleobase to the phosphodiamide as shown inFormula I and may have one or more additional chiral centers dependingon substituent selection. For example, each of the compounds of Examples1-28B herein also has an asymmetric phosphorus center. Accordingly, acompound of Formula I may have multiple chiral centers (also referred toas asymmetric or stereogenic centers). This invention encompassescompounds having either the (R) or (S) stereo-configuration at aphosphorus asymmetric center and at any additional asymmetric centersthat may be present in a compound of Formula I, as well asstereo-isomeric mixtures thereof.

This invention includes individual diastereomers, particularly epimers,i.e., compounds having the same chemical formula but which differ in thespatial arrangement around a single atom. This invention also includesmixtures of diastereomers, particularly mixtures of epimers, in allratios. Embodiments of this invention also include a mixture of epimersenriched with 51% or more of one of the epimers, including for example60% or more, 70% or more, 80% or more, or 90% or more of one epimer. Asingle epimer is preferred. An individual or single epimer refers to anepimer obtained by chiral synthesis and/or using generally knownseparation and purification techniques, and which may be 100% of oneepimer or may contain small amounts (e.g., 10% or less) of the oppositeepimer. Thus, individual diasteromers are a subject of the invention inpure form, both as levorotatory and as dextrorotatory antipodes, in theform of racemates and in the form of mixtures of the two diasteromers inall ratios. In the case of a cis/trans isomerism the invention includesboth the cis form and the trans form as well as mixtures of these formsin all ratios.

The preparation of individual stereoisomers can be carried out, ifdesired, by separation of a mixture by customary methods, for example bychromatography or crystallization, by the use of stereochemicallyuniform starting materials for the synthesis or by stereoselectivesynthesis. Optionally a derivatization can be carried out before aseparation of stereoisomers. The separation of a mixture ofstereoisomers can be carried out at an intermediate step during thesynthesis of a compound of Formula I or it can be done on a finalracemic product. Absolute stereochemistry may be determined by X-raycrystallography of crystalline products or crystalline intermediateswhich are derivatized, if necessary, with a reagent containing astereogenic center of known configuration. Alternatively, absolutestereochemistry may be determined by Vibrational Circular Dichroism(VCD) spectroscopy analysis. The present invention includes all suchisomers, as well as salts, solvates (which includes hydrates) andsolvated salts of such racemates, enantiomers, diastereomers andtautomers and mixtures thereof.

The atoms in a compound of Formula I may exhibit their natural isotopicabundances, or one or more of the atoms may be artificially enriched ina particular isotope having the same atomic number, but an atomic massor mass number different from the atomic mass or mass numberpredominantly found in nature. The present invention is meant to includeall suitable isotopic variations of the compounds of Formula I; forexample, different isotopic forms of hydrogen (H) include protium (¹H)and deuterium (²H). Protium is the predominant hydrogen isotope found innature. Enriching for deuterium may afford certain therapeuticadvantages, such as increasing in vivo half-life or reducing dosagerequirements, or may provide a compound useful as a standard forcharacterization of biological samples. Isotopically-enriched compoundsof Formula I can he prepared without undue experimentation byconventional techniques well known to those skilled in the art or byprocesses analogous to those described in the Schemes and Examplesherein using appropriate isotopically-enriched reagents and/orintermediates.

The compounds can be administered in the form of pharmaceuticallyacceptable salts. The term “pharmaceutically acceptable salt” refers toa salt which is not biologically or otherwise undesirable (e.g., isneither toxic nor otherwise deleterious to the recipient thereof) Sincethe compounds of Formula I contain by definition at least one basicgroup, the invention includes the corresponding pharmaceuticallyacceptable salts. When the compounds of Formula I contain one or moreacidic groups, the invention also includes the correspondingpharmaceutically acceptable salts. Thus, the compounds of Formula I thatcontain acidic groups (e.g., —COOH) can he used according to theinvention as, for example but not limited to, alkali metal salts,alkaline earth metal salts or as ammonium salts. Examples of such saltsinclude but are not limited to sodium salts, potassium salts, calciumsalts, magnesium salts or salts with ammonia or organic amines such as,for example, ethylamine, ethanolamine, triethanolamine or amino acids.Compounds of Formula I, which contain one or more basic groups, i.e.groups which can be protonated, can be used according to the inventionin the form of their acid addition salts with inorganic or organic acidsas, for example but not limited to, salts with hydrogen chloride,hydrogen bromide, phosphoric acid, sulfuric acid, nitric acid,benzenesulfonic acid, methanesulfonic acid, p-toluenesulfonic acid,naphthalenedisulfonic acids, oxalic acid, acetic acid, trifluoroaceticacid, tartaric acid, lactic acid, salicylic acid, benzoic acid, formicacid, propionic acid, pivalic acid, diethylacetic acid, malonic acid,succinic acid, pimelic acid, fumaric acid, maleic acid, malic acid,sulfaminic acid, phenylpropionic acid, gluconic acid, ascorbic acid,isonicotinic acid, citric acid, adipic acid, etc. If the compounds ofFormula I simultaneously contain acidic and basic groups in the moleculethe invention also includes, in addition to the salt forms mentioned,inner salts or betaines (zwitterions). Salts can be obtained from thecompounds of Formula I by customary methods which are known to theperson skilled in the art, for example by combination with an organic orinorganic acid or base in a solvent or dispersant, or by anion exchangeor cation exchange from other salts. The present invention also includesall salts of the compounds of Formula I which, owing to lowphysiological compatibility, are not directly suitable for use inpharmaceuticals but which can be used, for example, as intermediates forchemical reactions or for the preparation of pharmaceutically acceptablesalts.

The instant invention encompasses any composition comprised of acompound of Formula I or a compound that is a salt thereof, includingfor example but not limited to, a composition comprised of said compoundassociated together with one or more additional molecular and/or ioniccomponent(s) which may be referred to as a “co-crystal.” The term“co-crystal” as used herein refers to a solid phase (which may or maynot be crystalline) wherein two or more different molecular and/or ioniccomponents (generally in a stoichiometric ratio) are held together bynon-ionic interactions including but not limited to hydrogen-bonding,dipole-dipole interactions, dipole-quadrupole interactions or dispersionforces (van der Waals). There is no proton transfer between thedissimilar components and the solid phase is neither a simple salt nor asolvate. A discussion of co-crystals can be found, e.g., in S.Aitipamula et al., Crystal Growth and Design, 2012, 12 (5), pp.2147-2152.

More specifically with reference to this invention, a co-crystal iscomprised of a compound of Formula I or a pharmaceutically acceptablesalt thereof, and one or more non-pharmaceutically active component(s)which is not biologically or otherwise undesirable (e.g., is neithertoxic nor otherwise deleterious to the recipient thereof). Co-crystalscan be obtained from a compound of Formula I, or a pharmaceuticallyacceptable salt thereof, by customary methods known in the chemicalarts. For example, co-crystals comprised of a compound of this inventioncould be prepared by adding an acid or a neutral molecule at the desiredstoichiometry to the compound, adding an appropriate solvent to achievedissolution and, for example, precipitating, lyophilizing orconcentrating the solution to obtain the solid composition. Theco-crystal can be, but is not limited to, an embodiment wherein thecomposition is comprised of a neutral compound (i.e. not a salt form) ofFormula I and one or more non-pharmaceutically active component(s) andin a further embodiment, the co-crystal composition is crystalline.Crystalline compositions may be prepared, for example, by adding an acidor a neutral molecule at the desired stoichiometry to the compound ofFormula I, adding an appropriate solvent and heating to achieve completedissolution, and then allowing the solution to cool and the crystals togrow. The present invention also includes all co-crystals of thecompounds of this invention which, owing to low physiologicalcompatibility, are not directly suitable for use in pharmaceuticals butwhich can be used, for example, as intermediates for chemical reactionsor for the preparation of pharmaceutically acceptable co-crystals orsalts.

Furthermore, compounds of the present invention may exist in amorphousform and/or one or more crystalline forms, and as such all amorphous andcrystalline forms and mixtures thereof of the compounds of Formula I andsalts thereof are intended to be included within the scope of thepresent invention, In addition, some of the compounds of the instantinvention may form solvates with water (i.e., a hydrate) or commonorganic solvents. Such solvates and hydrates, particularly thepharmaceutically acceptable solvates and hydrates, of the compounds ofthis invention are likewise encompassed within the scope of thecompounds defined by Formula I and the pharmaceutically acceptable saltsthereof, along with un-solvated and anhydrous forms of such compounds.

Accordingly, the compounds of Formula I, embodiments thereof andspecific compounds described and claimed herein encompass all possiblesalts, including pharmaceutically acceptable salts, stereoisomers,tautomers, physical forms (e.g., amorphous and crystalline forms),co-crystal forms, solvate and hydrate forms, and any combination of theforegoing forms where such forms are possible.

The compounds of Formula I (or any embodiment thereof includingpharmaceutically acceptable salts thereof) are prodrugs. A discussion ofprodrugs is provided in (a) Stella, V. J.; Borchardt, R. T.; Hageman, M.J.; Oliyai, R.; Mag, H. et al. Prodrugs: Challenges and Rewards Part 1and Part 2; Springer, p. 726: New York, NY, USA, 2007, (b) Rautio,Kutnpulainen, Heimbach, T,; Oliyai, R.; Oh, D. et al. Prodrugs: designand clinical applications. Nat. Rev. Drug Discov. 2008, 7, 255, (c) T.Higuchi and V. Stella, Pro-drugs as Novel Delivery Systems (1987) 14 ofthe A.C.S. Symposium Series, and in (d) Bioreversible Carriers in DrugDesign, (1987) Edward B. Roche, ed., American Pharmaceutical Associationand Pergamon Press. More specifically, compounds of Formula I (or anyembodiment thereof and pharmaceutically acceptable salts thereof) areprodrug modifications of tenofovir, which is a mono-phosphonate. Thecompounds of Formula I may be converted intracellularly (in vivo or invitro) to the corresponding monophosphate or diphosphate of tenofovir.The conversion may occur by one or more mechanisms, e.g., anenzyme-catalyzed chemical reaction, a metabolic chemical reaction,and/or a spontaneous chemical reaction (e.g., solvolysis), such as, forexample, through hydrolysis in blood. While not wishing to be bound byany particular theory, tenofovir diphosphate is generally understood tobe responsible for inhibiting the HIV RT enzyme and for the resultingantiviral activity after administration of the compound of Formula I toa subject,

Another embodiment of the present invention is a compound of Formula Iwherein the compound or its salt is in a substantially pure form. Asused herein “substantially pure” means suitably at least about 60 wt. %,typically at least about 70 wt. %, preferably at least about 80 wt. %,more preferably at least about 90 wt. % (e.g., from about 90 wt. % toabout 99 wt. %), even more preferably at least about 95 wt. % (e.g.,from about 95 wt. % to about 99 wt. % or from about 98 wt. % to 100 wt.%), and most preferably at least about 99 wt. % (e.g. 100 wt. %) of aproduct containing a compound of Formula I or its salt (e.g., theproduct isolated from a reaction mixture affording the compound or salt)consists of the compound or salt. The level of purity of the compoundsand salts can be determined using a standard method of analysis such as,high performance liquid chromatography, and/or mass spectrometry or NMRtechniques. If more than one method of analysis is employed and themethods provide experimentally significant differences in the level ofpurity determined, then the method providing the highest purity levelgoverns. A compound or salt of 100% purity is one which is free ofdetectable impurities as determined by a standard method of analysis.With respect to a compound of Formula I which has one or more asymmetriccenters and can occur as mixtures of stereoisomers, a substantially purecompound can be either a substantially pure mixture of the stereoisomersor a substantially pure individual stereoisomer,

The compounds of Formula I herein, and pharmaceutically acceptable saltsthereof, are useful for HIV reverse transcriptase inhibition and forinhibiting HIV replication in vitro and in vivo. More particularly, thecompounds of Formula I for inhibiting the polymerase function of HIV-1reverse transcriptase. The testing of compounds of the Examples of thisinvention in the Viking assay set forth in Example 29 below, illustratethe ability of compounds of the invention to inhibit the RNA-dependentDNA polymerase activity of HIV-1 reverse transcriptase. The compounds ofFormula I may also be useful agents against HIV-2. The compounds ofExamples 1-28 (inclusive of A and B isomers) of the present inventionalso exhibit activity against drug resistant forms of HIV (e.g.,NNRTI-associated mutant strains K103N and/or Y181C; NRTI-associatedmutant strains M184V and M184I mutants).

This invention also encompasses methods for the treatment or prophylaxisof infection by HIV, for the inhibition of HIV reverse transcriptase,for the treatment, prophylaxis, or delay in the onset of AIDS in asubject in need thereof, which comprise administering to the subject aneffective amount of a compound of the invention or a pharmaceuticallyacceptable salt thereof.

The invention further encompasses methods for the treatment orprophylaxis of infection by HIV, for the inhibition of HIV reversetranscriptase, for the treatment, prophylaxis, or delay in the onset ofAIDS in a subject in need thereof, which comprise administering to thesubject an effective amount of a compound of the invention or apharmaceutically acceptable salt thereof in combination with aneffective amount of one or more additional anti-HIV agents selected fromthe group consisting of HIV antiviral agents, immunomodulators, andanti-infective agents. Within this embodiment, the anti-HIV agent is anantiviral selected from the group consisting of HIV protease inhibitors,HIV reverse transcriptase inhibitors, HIV integrase inhibitors, HIVfusion inhibitors, HIV entry inhibitors, and. HIV maturation inhibitors.

The invention encompasses a pharmaceutical composition comprising aneffective amount of a compound of the invention, or a pharmaceuticallyacceptable salt thereof, and a pharmaceutically acceptable carrier, Theinvention also encompasses a pharmaceutical composition comprising aneffective amount of a compound of the invention, or a pharmaceuticallyacceptable salt thereof, and a pharmaceutically acceptable carrierfurther comprising an effective amount of one or more additionalanti-HIV agents selected from the group consisting of HIV antiviralagents, immunomodulators, and anti-infective agents. Within thisembodiment, the anti-HIV agent is an antiviral selected from the groupconsisting of HIV protease inhibitors, HIV reverse transcriptaseinhibitors, HIV integrase inhibitors, HIV fusion inhibitors, HIV entryinhibitors, and HIV maturation inhibitors.

The compounds of this invention could also be useful for inhibition ofhepatitis B virus (HBV) reverse transcriptase. Accordingly, thisinvention also encompasses methods for the treatment of chronichepatitis B infection which comprise administering to the subject aneffective amount of a compound of the invention or a pharmaceuticallyacceptable salt thereof

The invention also encompasses a compound of the invention, or apharmaceutically acceptable salt thereof, for use in the preparation ofa medicament for the treatment or prophylaxis of infection by HIV, forthe inhibition of HIV reverse transcriptase, or for the treatment,prophylaxis, or delay in the onset of AIDS in a subject in need thereof

Other embodiments of the present invention include the following:

(a) A pharmaceutical composition comprising an effective amount of acompound of Formula I or a pharmaceutically acceptable salt thereof, anda pharmaceutically acceptable carrier. In particular embodiments, thepharmaceutical composition comprises a therapeutically effective amountof a compound of Formula I or a pharmaceutically acceptable saltthereof, and a pharmaceutically acceptable carrier. In otherembodiments, the pharmaceutical composition comprises a prophylacticallyeffective amount of a compound of Formula I or a pharmaceuticallyacceptable salt thereof, and a pharmaceutically acceptable carrier.

(b) A pharmaceutical composition which comprises the product prepared bycombining (e.g., mixing) an effective amount of a compound of Formula Ior a pharmaceutically acceptable salt thereof, and a pharmaceuticallyacceptable carrier.

(c) The pharmaceutical composition of (a) or (b), further comprising aneffective amount of one or more an anti-HIV agents selected from thegroup consisting of HIV antiviral agents, immunomodulators, andanti-infective agents.

(d) The pharmaceutical composition of (c), wherein the anti-HIV agent isselected from one or more of an antiviral selected from the groupconsisting of HIV protease inhibitors, nucleoside HIV reversetranscriptase inhibitors, non-nucleoside HIV reverse transcriptaseinhibitors, HIV integrase inhibitors, HIV fusion inhibitors. HIV entryinhibitors and HIV maturation inhibitors.

(e) A combination which is (i) a compound of Formula I or apharmaceutically acceptable salt thereof and (ii) an anti-HIV agentselected from the group consisting of HIV antiviral agents,immunomodulators, and anti-infective agents; wherein the compound andthe anti-HIV agent are each employed in an amount that renders thecombination effective for inhibition of HIV reverse transcriptase, fortreatment or prophylaxis of infection by HIV, or for treatment,prophylaxis of or delay in the onset or progression of AIDS.

(f) The combination of (e), wherein the anti-HIV agent is an antiviralselected from the group consisting of HIV protease inhibitors.nucleoside HIV reverse transcriptase inhibitors, non-nucleoside HIVreverse transcriptase inhibitors, HIV integrase inhibitors, HIV fusioninhibitors, HIV entry inhibitors and HIV maturation inhibitors.

(g) A method for the inhibition of HIV reverse transcriptase in asubject in need thereof which comprises administering to the subject aneffective amount of a compound of Formula I or pharmaceuticallyacceptable salt thereof. In specific embodiments, the method comprisesadministering to the subject a therapeutically effective amount of acompound of Formula I or a pharmaceutically acceptable salt thereof. Inother embodiments, the method comprises administering to the subject aprophylactically effective amount of a compound of Formula I or apharmaceutically acceptable salt thereof.

(h) A method for the prophylaxis or treatment of infection by HIV (e.g.,HIV-1) in a subject in need thereof which comprises administering to thesubject an effective amount of a compound of Formula I orpharmaceutically acceptable salt thereof. In specific embodiments, themethod comprises administering to the subject a therapeuticallyeffective amount of a compound of Formula I or a pharmaceuticallyacceptable salt thereof. In other embodiments, the method comprisesadministering to the subject a prophylactically effective amount of acompound of Formula I or a pharmaceutically acceptable salt thereof.

(i) The method of (h), wherein the compound of Formula I or apharmaceutically acceptable salt thereof is administered in combinationwith an effective amount of at least one other HIV antiviral selectedfrom the group consisting of RN protease inhibitors, integraseinhibitors, non-nucleoside HIV reverse transcriptase inhibitors,nucleoside HIV reverse transcriptase inhibitors, HIV fusion inhibitors,HIV entry inhibitors and HIV maturation inhibitors.

(j) A method for the prophylaxis, treatment or delay in the onset orprogression of AIDS in a subject in need thereof which comprisesadministering to the subject an effective amount of a compound ofFormula I or pharmaceutically acceptable salt thereof. In specificembodiments, the method comprises administering to the subject atherapeutically effective amount of a compound of Formula I or apharmaceutically acceptable salt thereof. In other embodiments, themethod comprises administering to the subject a prophylacticallyeffective amount of a compound of Formula I or a pharmaceuticallyacceptable salt thereof

(k) The method of (j), wherein the compound is administered incombination with an effective amount of at least one other HIV antiviralselected from the group consisting of HIV protease inhibitors, HIVintegrase inhibitors, non-nucleoside HIV reverse transcriptaseinhibitors, nucleoside HIV reverse transcriptase inhibitors, HIV fusioninhibitors, HIV entry inhibitors and HIV maturation inhibitors.

(l) A method for the inhibition of HIV reverse transcriptase in asubject in need thereof which comprises administering to the subject thepharmaceutical composition of (a), (b), (c) or (d) or the combination of(e) or (f).

(m) A method for the prophylaxis or treatment of infection by HIV (e.g.,HIV-1) in a subject in need thereof, which comprises administering tothe subject the pharmaceutical composition of (a), (b), (c) or (d) orthe combination of (e) or (f).

(n) A method for the prophylaxis, treatment, or delay in the onset orprogression of AIDS in a subject in need thereof which comprisesadministering to the subject the pharmaceutical composition of (a), (b),(c) or (d) or the combination of (e) or (f).

The present invention also includes a compound of Formula I orpharmaceutically acceptable salt thereof (i) for use in, (ii) for use asa medicament for, or (iii) for use in the preparation of a medicamentfor: (a) therapy (e.g., of the human body), (b) medicine, (c) inhibitionof HIV reverse transcriptase, (d) treatment or prophylaxis of infectionby HIV, or (e) treatment, prophylaxis of, or delay in the onset orprogression of AIDS. In these uses, the compounds of the presentinvention can optionally be employed in combination with one or moreanti-HIV agents selected from HIV antiviral agents, anti-infectiveagents, and immunomodulators,

Additional embodiments of the invention include the pharmaceuticalcompositions, combinations and methods set forth in (a)-(n) above andthe uses (i)(a)-(e) through (iii)(a)-(e) set forth in the precedingparagraph, wherein the compound of the present invention employedtherein is a compound of one of the embodiments, aspects, classes,sub-classes, or features described above. In all of these embodimentsetc., the compound may optionally be used in the form of apharmaceutically acceptable salt.

Additional embodiments of the present invention include each of thecompounds, pharmaceutical compositions, combinations, methods and usesset forth in the preceding paragraphs, wherein the compound or its saltemployed therein is substantially pure. With respect to a pharmaceuticalcomposition comprising a compound of Formula I or its salt and apharmaceutically acceptable carrier and optionally one or moreexcipients, it is understood that the term “substantially pure” is inreference to a compound of Formula I or its salt per se.

Still additional embodiments of the present invention include thepharmaceutical compositions, combinations and methods set forth in(a)-(n) above and the uses (i)(a)-(e) through (iii)(a)-(e) set forthabove, wherein the HIV of interest is HIV-1. Thus, for example, in thepharmaceutical composition (d), the compound of Formula. I is employedin an amount effective against HIV-1 and the anti-HIV agent is an HIV-1antiviral selected from the group consisting of HIV-1 proteaseinhibitors, HIV-1 reverse transcriptase inhibitors, HIV-1 integraseinhibitors, HIV-1 fusion inhibitors, HIV-1 entry inhibitors and HIV-1maturation inhibitors. The compounds of Formula I may also be usefulagents against HIV-2.

The term “administration” and variants thereof (e.g., “administering” acompound) in reference to a compound of Formula I means providing thecompound to the individual in need of treatment or prophylaxis andincludes both self-administration and administration to the patient byanother person. When a compound is provided in combination with one ormore other active agents (e.g., antiviral agents useful for treating orprophylaxis of HIV infection or AIDS), “administration” and its variantsare each understood to include provision of the compound and otheragents at the same time or at different times. When the agents of acombination are administered at the same time, they can be administeredtogether in a single composition or they can be administered separately.

As used herein, the term “composition” is intended to encompass aproduct comprising the specified ingredients, as well as any productwhich results from combining the specified ingredients. Ingredientssuitable for inclusion in a pharmaceutical composition arepharmaceutically acceptable ingredients, which means the ingredientsmust be compatible with each other and not deleterious to the recipientthereof.

The term “subject” or “patient” as used herein refers to an animal,preferably a mammal, most preferably a human, who has been the object oftreatment, observation or experiment.

The term “effective amount” as used herein means an amount of a compoundsufficient to inhibit HIV reverse transcriptase, inhibit HIVreplication, exert a prophylactic effect, and/or a exert a therapeuticeffect after administration. One embodiment of “effective amount” is a“therapeutically effective amount” which is an amount of a compound thatis effective for inhibiting HIV reverse transcriptase, inhibiting HIVreplication (either of the foregoing which may also be referred toherein as an “inhibition effective amount”), treating HIV infection,treating AIDS, delaying the onset of AIDS, and/or slowing progression ofARC or AIDS in a patient infected with HIV. Another embodiment of“effective amount” is a “prophylactically effective amount” which is anamount of the compound that is effective for prophylaxis of HIVinfection in a subject not infected with HIV or prophylaxis of ARC orAIDS in an HIV-infected patient. It is understood that an effectiveamount can simultaneously be both a therapeutically effective amount,e.g., for treatment of HIV infection, and a prophylactically effectiveamount, e.g., for prevention or reduction of risk for developing AIDS ina subject infected with HIV. The term “preventing,” as used herein withrespect to an HIV viral infection or AIDS, refers to reducing thelikelihood or severity of HIV infection or AIDS. When the compound ofFormula I is administered as a salt, reference to an amount of thecompound in milligrams or grams is based on the free form (i.e., thenon-salt form) of the compound. In the combination therapies of thepresent invention, an effective amount can refer to each individualagent or to the combination as a whole, wherein the amounts of allagents administered in the combination are together effective, butwherein a component agent of the combination may or may not be presentindividually in an effective amount with reference to what is consideredeffective for that component agent if it were administered alone.

In the method of the present invention (i.e., inhibiting HIV reversetranscriptase, treating or prophylaxis of HIV infection, inhibiting HIVreplication, treating or prophylaxis of AIDS, delaying the onset ofAIDS, or delaying or slowing progression of AIDS), the compounds of thisinvention, optionally in the form of a. salt, can be administered bymeans that produce contact of the active agent with the agent's site ofaction. They can be administered by conventional means available for usein conjunction with pharmaceuticals, either as individual therapeuticagents or in a combination of therapeutic agents. They can beadministered alone, but typically are administered with a pharmaceuticalcarrier selected on the basis of the chosen route of administration andstandard pharmaceutical practice. The compounds of the invention can,for example, be administered orally (e.g., via tablet or capsule),parenterally (including subcutaneous injections, intravenous,intramuscular or intrasternal injection, or infusion techniques), byinhalation spray, or rectally, in the form of a unit dosage of apharmaceutical composition containing an effective amount of thecompound and conventional non-toxic pharmaceutically acceptablecarriers, adjuvants and vehicles. The compound could also beadministered via an implantable drug delivery device adapted to providean effective amount of the compound or a pharmaceutical composition ofthe compound over an extended period of time, for example but notlimited to, over the course of a month, 3 months, 6 months or a year.

Solid preparations suitable for oral administration (e.g., powders,pills, capsules and tablets) can be prepared according to techniquesknown in the art and can employ such solid excipients as starches,sugars, kaolin, lubricants, binders, disintegrating agents and the like.Liquid preparations suitable for oral administration (e.g., suspensions,syrups, elixirs and the like) can be prepared according to techniquesknown in the art and can employ any of the usual media such as water,glycols, oils, alcohols and the like. Parenteral compositions can beprepared according to techniques known in the art and typically employsterile water as a carrier and optionally other ingredients, such as asolubility aid. Injectable solutions can be prepared according tomethods known in the art wherein the carrier comprises a salinesolution, a glucose solution or a solution containing a mixture ofsaline and glucose. Implantable compositions can be prepared accordingto methods known in the art wherein the carrier comprises the activechemical ingredient with polymers as suitable excipients, or utilizingan implantable device for drug delivery. Further description of methodssuitable for use in preparing pharmaceutical compositions for use in thepresent invention and of ingredients suitable for use in saidcompositions is provided in Remington's Pharmaceutical Sciences, 18^(th)edition, edited by A. R. Gennaro, Mack Publishing Co., 1990 and inRemington—The Science and Practice of Pharmacy, 22nd Edition, publishedby Pharmaceutical Press and Philadelphia College of Pharmacy atUniversity of the Sciences, 2012, ISBN 978 0 85711-062-6 and prioreditions.

Formulations of compounds described by Formula. I that result in drugsupersaturation and/or rapid dissolution may be utilized to facilitateoral drug absorption. Formulation approaches to cause drugsupersaturation and/or rapid dissolution include, but are not limitedto, nanoparticulate systems, amorphous systems, solid solutions, soliddispersions, and lipid systems. Such formulation approaches andtechniques for preparing them are known in the art. For example, soliddispersions can be prepared using excipients and processes as describedin reviews (e.g., A.T.M. Serajuddin, J Pharm Sci, 88:10, pp. 1058-1066(1999)). Nanoparticulate systems based on both attrition and directsynthesis have also been described in reviews such as Wu et al (F.Kesiso_(L)tiou, S. Panmai, Y. Wu, Advanced Drug Delivery Reviews, 59:7pp. 631-644 (2007)).

The compounds of Formula I can be administered in a dosage range of0.001 to 1000 mg/kg of mammal (e.g., human) body weight per day, or atlonger time intervals on non-consecutive days as appropriate, in asingle dose or in divided doses. One example of a dosage range is 0.01to 500 mg/kg body weight per day, or at other time intervals asappropriate, administered orally or via other routes of administrationin a single dose or in divided doses. Another example of a dosage rangeis 0.1 to 100 mg/kg body weight per day, or at other time intervals asappropriate, administered orally or via other routes of administrationin single or divided doses. Another example of a dosage range is 50 mgto 1 gram per day, in a single dose or divided doses.

Daily or weekly administration or less frequent dosing regimens withlonger time intervals on non-consecutive days (as discussed below), canbe via any suitable route of administration, e.g., but not limited to,oral or parenteral. Daily or weekly administration is preferably viaoral administration. For either a daily or weekly dosing regimen, oneach day (calendar day or about a 24 hour period of time) of drugadministration (the “administration day”), the desired dosage amount maybe administered once per administration day or in divided dosage amountsadministered at two or more staggered times during the administrationday, e.g., a first administration followed about 12 hours later with asecond administration during the course of an administration day (the“dosage times)”). The desired dosage amount at each of the one or moredosage times on an administration day can be administered via one oraldosage unit such as a tablet, or more than one oral dosage unit asappropriate. Preferably the administration is via a single oral dosageunit, e.g, a tablet, once per administration day.

For weekly or less frequent dosing regimens with longer time intervalson non-consecutive days, a parenteral route of administration may beemployed. Examples of dosing regimens with longer time intervals onnon-consecutive days include but are not limited to administrationweekly (every seventh day with leeway as to exact date of dosing),bi-weekly (every two weeks with leeway as to exact date of dosing),monthly (e.g., every 30 days, or the same calendar day each month withleeway as to exact date of dosing), bimonthly (e.g., every 60 days, orthe same calendar day every two months with leeway as to exact date ofdosing), every 3 months (e.g., every 90 days, or the same calendar dayevery three months with leeway as to exact date of dosing), every sixmonths (e.g., every 180 days, or the same calendar day every six monthswith leeway as to exact date of dosing), or yearly (e.g., every 12months with leeway as to exact date of the annual dosing). “Leeway” isintended to mean that the dosing regimens described herein alsoencompasses those wherein the patient generally follows the timeintervals between administration days including when the interval is notalways strictly followed by the patient, e.g., a weekly dosing regimenwhere the patient may take the drug product the day before or the dayafter the seventh day following the prior administration day for one ormore weeks. The leeway time may increase as the dosing regimen intervalincreases. For oral (e.g., tablets or capsules) or other routes ofadministration, the dosage units may contain 1.0 mg to 1000 mg of theactive ingredient, for example but not limited to, 1, 5, 10, 15, 20, 25,50, 75, 100, 150, 200, 250, 300, 350, 400, 450, 500, 600, 700, 800, 900or 1000 milligrams of the active ingredient for the symptomaticadjustment of the dosage to the patient to be treated. Furthermore, thecompound may be formulated in oral formulations for immediate ormodified release such as extended or controlled release.

The favorable pharmacokinetic profile of tested compounds of thisinvention may also render the compounds suitable for less frequentdosing. Thus, the compounds of the invention could be administeredorally, weekly or parenterally at longer time intervals as describedabove, For parenteral administration, the compositions can beadministered, e.g., intravenously (IV) or intramuscularly (IM) viainjection, or using other infusion techniques. One or more of suchinjections or infusions may be administered at each dosing time intervalas needed to deliver the appropriate amount of active agent. Thecompound could also be administered subcutaneously using an implantabledevice. For parenteral administration including implantable devicesemploying longer duration dosing intervals such as monthly, every 3months, every 6 months, yearly or longer intervals, the dosage amountwould be adjusted upward as needed to provide effective treatment duringthe time intervals between administration of each dose.

The specific dose level and frequency of dosage for any particularpatient may be varied and will depend upon a variety of factorsincluding the activity of the specific compound employed, the metabolicstability and length of action of that compound, the age, body weight,general health, sex, diet, mode and time of administration, rate ofexcretion, drug combination, the severity of the particular condition,and the host undergoing therapy. In sonic cases, depending on thepotency of the compound or the individual response, it may be necessaryto deviate upwards or downwards from the given dose. The amount andfrequency of administration will be regulated according to the judgmentof the attending clinician considering such factors.

As noted above, the present invention is also directed to use of acompound of Formula I with one or more anti-HIV agents. An “anti-HIVagent” is any agent which is directly or indirectly effective in theinhibition of HIV, the treatment or prophylaxis of HIV infection, and/orthe treatment, prophylaxis or delay in the onset or progression of AIDS.It is understood that an anti-HIV agent is effective in treating,preventing, or delaying the onset or progression of HIV infection orAIDS and/or diseases or conditions arising therefrom or associatedtherewith. For example, the compounds of this invention may beeffectively administered, whether at periods of pre-exposure and/orpost-exposure, in combination with effective amounts of one or moreanti-HIV agents selected from HIV antiviral agents, imunomodulators,antiinfectives, or vaccines useful for treating HIV infection or AIDS.Suitable HIV antivirals for use in combination with the compounds of thepresent invention include, for example, those listed in Table A asfollows, or a pharmaceutically acceptable salt thereof:

TABLE A Antiviral Agents for Treating HIV infection or AIDS Name Typeabacavir, ABC, Ziagen ® nRTI abacavir + lamivudine, Epzicom ® nRTIabacavir + lamivudine + zidovudine, Trizivir ® nRTI amprenavir,Agenerase ® PI atazanavir, Reyataz ® PI AZT, zidovudine, azidothymidine,Retrovir ® nRTI capravirine nnRTI darunavir, Prezista ® PI ddC,zalcitabine, dideoxycytidine, Hivid ® nRTI ddI, didanosine,dideoxyinosine, Videx ® nRTI ddI (enteric coated), Videx EC ® nRTIdelavirdine, DLV, Rescriptor ® nnRT1 dolutegravir, Tivicay ® InIdoravirine, MK-1439 nnRTI efavirenz, EFV, Sustiva ®, Stocrin ® nnRTIefavirenz + emtricitabine + tenofovir DF, Atripla ® nnRTI + nRTI EFdA(4′-ethynyl-2-fluoro-2′-deoxyadenosine) nRTI Elvitegravir InIemtricitabine, FTC, Emtriva ® nRTI emtricitabine + tenofovir DF,Truvada ® nRTI emvirine, Coactinon ® nnRTI enfuvirtide, Fuzeon ® FIenteric coated didanosine, Videx EC ® nRTI etravirine, TMC-125 nnRTIfosamprenavir calcium, Lexiva ® PI indinavir, Crixivan ® PI lamivudine,3TC, Epivir ® nRTI lamivudine + zidovudine, Combivir ® nRTI lopinavir PIlopinavir + ritonavir, Kaletra ® PI maraviroc, Selzentry ® EInelfinavir, Viracept ® PI nevirapine, NVP, Viramune ® nnRTI PPL-100(also known as PL-462) (Ambrilia) PI raltegravir, MK-0518, Isentress ™InI Rilpivirine nnRTI ritonavir, Norvir ® PI saquinavir, Invirase ®,Fortovase ® PI stavudine, d4T, didehydrodeoxythymidine, Zerit ® nRTItipranavir, Aptivus ® PI vicriviroc EI EI = entry inhibitor; FI = fusioninhibitor; InI = integrase inhibitor; PI = protease inhibitor; nRTI =nucleoside reverse transcriptase inhibitor; nnRTI = non-nucleosidereverse transcriptase inhibitor. Some of the drugs listed in the tableare used in a salt form; e.g., abacavir sulfate, delavirdine mesylate,indinavir sulfate, atazanavir sulfate, nelfinavir mesylate, saquinavirmesylate.

It is understood that the scope of combinations of the compounds of thisinvention with anti-HIV agents is not limited to the HIV antiviralslisted in Table A, but includes in principle any combination with anypharmaceutical composition useful for the treatment or prophylaxis ofAIDS. The HIV antiviral agents and other agents will typically beemployed in these combinations in their conventional dosage ranges andregimens as reported in the art, including, for example, the dosagesdescribed in the Physicians' Desk Reference, Thomson PDR, Thomson PDR,57^(th) edition (2003), the 58^(th) edition (2004), or the 59^(th)edition (2005) and the Physicians' Desk Reference (68^(th) ed.). (2014),Montvale, N.J.: PDR Network. The dosage ranges for a compound of theinvention in these combinations can be the same as those set forthabove.

The compounds of this invention are also useful in the preparation andexecution of screening assays for antiviral compounds. For example, thecompounds of this invention can be useful for isolating enzyme mutants,which are excellent screening tools for more powerful antiviralcompounds. Furthermore, the compounds of this invention can be useful inestablishing or determining the binding site of other antivirals to HIVreverse transcriptase, e.g., by competitive inhibition.

Abbreviations and acronyms employed herein include the following:

Ac acetyl ACN acetonitrile AcOH or HOAc acetic acid APCIatmospheric-pressure chemical ionization Aq aqueous amu atomic massunits Bn benzyl Boc or BOC tert-butoxycarbonyl Bu butyl Bz benzoylcalc'd calculated cBu cyclobutyl Cbz benyzloxycarbonyl cHex cyclohexylcPen cyclopentyl cPr cyclopropyl DBU 1,8-diazabicycloundec-7-ene DCE1,2-dichloroethane DCM dichloromethane DIEA or Hünig'sN,N-diisopropylethylamine base DMA 1,2-dimethylacetamide DMFdimethylformamide DMSO dimethyl sulfoxide EDC 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide EDTA ethylenediamine tetraacetic acidESI electrospray ionization Et ethyl EtOH ethanol EtOAc ethyl acetate Ggrams H hour HATU N,N,N′,N′,-tetramethyl-O-(7- azabenzotriazol-1-yl)uronium hexafluorophosphate HOBt 1-hydroxybenzotriazole HPLChigh-performance liquid chromatography IPA isopropanol iPr isopropyl LCliquid chromatography LCMS liquid chromatography mass spectrometry mCPBAm-choroperoxybenzoic acid Me Methyl MeOH methanol mg milligrams minMinute μL microliters mL milliliters mm millimeter mmol millimoles μMmicromolar MS mass spectrometry Ms methanesulfonyl (mesyl) MTBE methyltert-butyl ether nm nanometer nM nanomolar NMR nuclear magneticresonance spectroscopy obsv'd observed Ph Phenyl Pr Propyl rac racemicmixture RT or rt room temperature (ambient, about 25° C.) sat or sat'd.saturated SFC supercritical fluid chromatography tBu tert-butyl TEAtriethylamine (Et₃N) TFA trifluoroacetic acid THF tetrahydrofuran TLCthin layer chromatographyMethods for preparing the exemplified compounds of this invention aredescribed in the following Scheme and Examples. Starting materials andintermediates were purchased commercially from common catalog sources orwere made using known procedures, or as otherwise illustrated. In somecases the order of carrying out the reaction steps in the schemes may bevaried to facilitate the reaction or to avoid unwanted reactionproducts.

Compounds of Formula IV can be prepared from(R)-(((1-(6-amino-9H-purin-9-yl)propan-2-yl)oxy)methyl)phosphonic acid(I, referred to herein as TFV) though a three-step sequence. First, TFVcan undergo functionalization with variably-substituted unnaturalgeminal amino esters and chlorophenol in a one-step one-pot condensationreaction with 2,2′-dipyridyldisulfide (Aldrithiol), triphenylphosphine,and base to provide intermediates of Formula II. This intermediate isthen hydrolyzed in the presence of base and water to provideintermediates of Formula III, which are unstable and are usedimmediately in the subsequent coupling reaction. Intermediates ofFormula III can then undergo functionalization with variably-substitutedunnatural D-amino esters by a condensation reaction with2,2′-dipyridyldisulfide (Aldrithiol), triphenylphosphine, and base toprovide intermediates of the current invention (Formula IV). Aminoesters that are not commercially available can be readily prepared bycondensation between the corresponding amino acid and alcohols withthionyl chloride.

Reactions sensitive to moisture or air were performed under nitrogen orargon using anhydrous solvents and reagents. The progress of reactionswas determined by either analytical thin layer chromatography (TLC)usually performed with E. Merck pre-coated TLC plates, silica gel60F-254, layer thickness 0.25 mm or liquid chromatography-massspectrometry (LC-MS).

Typically the analytical LC-MS system used consisted of a Waters ZQ™platform with electrospray ionization in positive ion detection modewith an Agilent 1100 series HPLC with autosampler, The column wascommonly a Waters Xterra MS C18, 3.0×50mm, 5 μm or a Waters AcquityUPLC® BEH C18 1.0×50 mm, 1.7 μm. The flow rate was 1 mL/min, and theinjection volume was 10 μL. UV detection was in the range 210-400 nm.The mobile phase consisted of solvent A (water plus 0.05% TFA) andsolvent B (acetonitrile plus 0.05% TFA) with a gradient of 100% solventA for 0.7 min changing to 100% solvent B over 3.75 min, maintained for1.1 min, then reverting to 100% solvent A over 0.2 mm.

Preparative HPLC purifications were usually performed using either amass spectrometry directed system or a non-mass spectrometry guidedsystem. Usually they were performed on a Waters ChromatographyWorkstation configured with a LC-MS System consisting of: Waters ZQ™single quad MS system with Electrospray Ionization, Waters 2525 GradientPump, Waters 2767 Injector/Collector, Waters 996 PDA Detector, the MSConditions of: 150-750 amu, Positive Electrospray, Collection Triggeredby MS, and a Waters SUNFIRE® C-18 5 micron, 30 mm (id)×100 mm column.The mobile phases consisted of mixtures of acetonitrile (10-100%) inwater containing 0.1% TEA. Flow rates were maintained at 50 mL/min, theinjection volume was 1800 μL, and the UV detection range was 210-400 nm.An alternate preparative HPLC system used was a Gilson Workstationconsisting of: Gilson GX-281 Injector/Collector, Gilson UV/VIS-155Detector, Gilson 333 and 334 Pumps, and either a Phenomenex Gemini-NXC-18 5 micron, 50 mm (id) x 250 mm column or a Waters XBridge™ C-18 5micron OBD™, 30 mm (id)×250 mm column. The mobile phases consisted ofmixtures of acetonitrile (0-75%) in water containing 5mmol (NH₄)HCO₃.Flow rates were maintained at 50 mL/min for the Waters Xbridge™ columnand 90 mL/min for the Phenomenex Gemini column. The injection volumeranged from 1000-8000 μL, and the UV detection range was 210-400 nm.Mobile phase gradients were optimized for the individual compounds.Reactions performed using microwave irradiation were normally carriedout using an Emrys Optimizer manufactured by Personal Chemistry, or anInitiator manufactured by Biotage. Concentration of solutions wascarried out on a rotary evaporator under reduced pressure. Flashchromatography was usually performed using either a Biotage® FlashChromatography apparatus (Dyax Corp.), an ISCO CombiFlash® Rf apparatus,or an ISCO CombiFlash® Companion XL on silica gel (32-63 μM, 60 Å poresize) in pre-packed cartridges of the size noted. ¹H NMR spectra wereacquired at 500 MHz spectrometers in CDCl₃ solutions unless otherwisenoted. Chemical shifts were reported in parts per million (ppm).Tetramethylsilane (TMS) was used as an internal reference in CD₃Clsolutions, and residual CH₃OH peak or TMS was used as an internalreference in CD₃OD solutions. Coupling constants (J) were reported inhertz (Hz). Chiral analytical chromatography was most commonly performedon one of CHIRALPAK® AS, CHIRALPAK® AD, CHIRALCEL® OD, CHIRALCEL® IA, orCHIRALCEL® OJ columns (250×4.6 mm) (Daicel Chemical Industries, Ltd.)with noted percentage of either ethanol in hexane (% Et/Hex) orisopropanol in heptane (% IPA/Hep) as isocratic solvent systems. Chiralpreparative chromatography was conducted on one of CHIRALPAK AS,CHIRALPAK AD, CHIRALCEL® OD, CHIRALCEL®IA, CHIRALCEL® OJ columns (20×250mm) (Daicel Chemical Industries, Ltd.) with desired isocratic solventsystems identified on chiral analytical chromatography or bysupercritical fluid (SFC) conditions.

Several methods for preparing the compounds of this invention are alsodescribed in the Examples. Starting materials and intermediates werepurchased commercially from common catalog sources or were made usingknown procedures, or as otherwise illustrated. Some frequently appliedroutes to the compounds of Formula I are also described by the schemesas illustrated in the following examples. In some cases the order ofcarrying out the steps of reaction schemes may be varied to facilitatethe reaction or to avoid unwanted reaction products.

It is understood that a chiral center in a compound may exist in the “S”or “R” stereoconfigurations, or as a mixture of both, unlessspecifically noted otherwise. In some of the following examples,compounds having a chiral center were separated into singlestereoisomers, referred to as Isomer A (faster eluting isomer) andIsomer B (slower eluting isomer), based on their observed elution orderresulting from the separation as performed. Except for the definedchiral centers in the parent stereoisomer mixture, absolutestereochemistry (R or S) of each of the separated stereoisomers was notdetermined, unless specifically noted otherwise. In examples whereabsolute stereochemistry of each of the separated isomers was notdetermined, an asterisk (*) may be used in the associated chemicalstructure drawing that indicates the location of the unassianed chiralcenter.

INTERMEDIATE A

(R)-isopropyl 2-aminopropanoate hydrochloride

A neat solution of isopropanol (51.6 ml, 673 mmol) was treated slowlywith thionyl chloride (9.01 ml, 123 mmol) at room temperature over a2-min period. The mixture was treated with D-alanine (10 g, 112 mmol)and fitted with a reflux condenser. This suspension was heated to 85° C.(reflux) and stirred overnight. The solution was concentrated todryness. Oily solids were suspended in hexanes and concentrated. Theresulting oil crystallizes upon standing and drying under high vacuum toprovide the title compound: LC/MS: [(M+1)]⁺=132.0.

INTERMEDIATE B

(R)-hexyl 2-aminopropanoate hydrochloride

A stirring solution of neat hexan-1-ol (3096 mg, 30.3 mmol) in a 40 mLscintillation vial was treated dropwise with thionyl chloride (406 μl,5.56 mmol) at room temperature over a 2 minute period. The mixture wastreated with D-alanine (500 mg, 5.61 mmol), capped and the suspensionwas heated to 80° C. and stirred overnight. The mixture was partitionedbetween dichloromethane (30 mL) and water (15 mL), The water layer waswashed with dichloromethane (30 mL) again and then concentrated,azeotroping twice with acetonitrile and dried under high vacuum toprovide the title compound: LC/MS: [(M+1)]⁺=174.1.

INTERMEDIATE C

(R)-pentyl 2-aminopropanoate hydrochloride

INTERMEDIATE C was prepared on a 4.85 mmol scale in a similar fashion tothat described for the synthesis of INTERMEDIATE B starting fromD-alanine, except using n-pentanol to provide the title compound: LC/MS:[(M+1)]⁺=160.1.

INTERMEDIATE D

(R)-butyl 2-aminopropanoate hydrochloride

INTERMEDIATE D was prepared on a 4.51 mmol scale in a similar fashion tothat described for the synthesis of INTERMEDIATE B starting fromD-alanine, except using n-butanol to provide the title compound: LC/MS:[(M+1)]⁺=146.0.

INTERMEDIATE E

(R)-isobutyl 2-aminopropanoate hydrochloride

INTERMEDIATE E was prepared on a 3.37 mmol scale in a similar fashion tothat described for the synthesis of INTERMEDIATE B starting fromD-alanine, except using isobutanol to provide the title compound: LC/MS:[(M+1)]⁺=146.0.

INTERMEDIATE F

(R)-cyclobutyl 2-aminopropanoate hydrochloride

INTERMEDIATE F was prepared on a 14.5 mmol scale in a similar fashion tothat described for the synthesis of INTERMEDIATE A starting fromD-alanine, except using cyclobutanol to provide the title compound:LC/MS: [(M+1)]⁺=144.0.

INTERMEDIATE G

(R)-cyclopentyl 2-aminonropanoate hydrochloride

INTERMEDIATE G was prepared on a 80 mmol scale in a similar fashion tothat described for the synthesis of INTERMEDIATE G starting fromD-alanine, except using cyclopentanol to provide the title compound:LC/MS: [(M+1)]⁺=158.0.

INTERMEDIATE H

(R)-cyclohexyl 2-aminopropanoate hydrochloride

INTERMEDIATE H was prepared on a 4 mmol scale in a similar fashion tothat described for the synthesis of INTERMEDIATE B starting fromD-alanine, except using cyclohexanol to provide the title compound:LC/MS: [(M+1)]⁺=172.1.

INTERMEDIATE I

isopropyl 2-amino-2-methylpropanoate hydrochloride

A neat solution of isopropanol (89 ml, 1164 mmol) was treated slowlywith thionyl chloride (15.57 ml, 213 mmol) at room temperature over a 2min period. The mixture was treated with 2-amino-2-methylpropanoic acid(20 g, 194 mmol) and fitted with a reflux condenser. This suspension washeated to 85° C. (reflux) and stirred for 3 days. The resulting clearsolution was concentrated to dryness. The resulting oil was crystallizedby trituration in diethyl ether and hexanes. The solids were isolated byfiltration and dried under high vacuum to provide the title compound:LC/MS: [(M+1)]⁺=146.1.

INTERMEDIATE J

butyl 2-amino-2-methylpropanoate hydrochloride

INTERMEDIATE J was prepared on a 8 mmol scale in a similar fashion tothat described for the synthesis of INTERMEDIATE I starting from2-amino-2-methylpropanoic acid, except using n-butanol to provide thetitle compound: LC/MS: [(M+1)]⁺=160.1.

INTERMEDIATE K

pentyl 2-amino-2-methylpropanoate hydrochloride

A neat solution of pentan-1-ol (205 g, 2327 mmol) was treated slowlywith thionyl chloride (15,57 ml, 213 mmol) at room temperature over a 10min period (exotherm to ˜60° C.). The mixture was treated with2-amino-2-methylpropanoic acid (40 g, 388 mmol) and fitted with a refluxcondenser and stirred at 80° C. over the weekend. The reaction mixturewas concentrated under reduced pressure and then dissolved in water (1L) and washed with 1:1 EtOAc/hexane (2×2 L), hexanes (1×2 L) anddichloromethane (3×1 L). The aqueous layer was concentrated andazeotroped with acetonitrile and then toluene. The residue was driedovernight under high vacuum. The solids were triturated with ˜500 mL ofdiethyl ether, filtered and dried to afford the title compound: LC/MS:[(M+1)]⁺=174.1.

INTERMEDIATE L

hexyl 2-amino-2-methylpropanoate hydrochloride

INTERMEDIATE L was prepared on a 388 mmol scale in a similar fashion tothat described for the synthesis of INTERMEDIATE K starting from2-amino-2-methylpropanoic acid, except using n-hexanol to provide thetitle compound: LC/MS: [(M+1)]⁺=188.1.

INTERMEDIATE M

isobutvl 2-amino-2-methylpropanoate hydrochloride

INTERMEDIATE M was prepared on a 9.15 mmol scale in a similar fashion tothat described for the synthesis of INTERMEDIATE K startimg from from2-amino-2-methylpropanoic acid, except using isobutanol to provide thetitle compound: LC/MS: [(M+1)]⁺=160.1.

INTERMEDIATE N

cyclohutyl 2-amino-2-methylpropanoate hydrochloride

INTERMEDIATE N was prepared on a 11.62 mmol scale in a similar fashionto that described for the synthesis of INTERMEDIATE K starting from from2-amino-2-methylpropanoic acid, except using cyclobutanol to provide thetitle compound: LC/MS: [(M+1)]⁺=158.0.

INTERMEDIATE O

cyclopentY1 2-amino-2-methylpropanoate hydrochloride

INTERMEDIATE O was prepared on a 15.84 mmol scale in a similar fashionto that described for the synthesis of INTERMEDIATE K starting from from2-amino-2-methylpropanoic acid, except using cyclopentanol to providethe title compound: LC/MS: [(M+1)]⁺=172.1.

INTERMEDIATE P

cyclohexyl 2-amino-2-methylproparioate hydrochloride

INTERMEDIATE P was prepared on a 9.2 mmol scale in a similar fashion tothat described for the synthesis of INTERMEDIATE K starting from from2-amino-2-methylpropanoic acid, except using cyclohexanol to provide thetitle compound: LC/MS: [(M+1)]⁺=186.1.

INTERMEDIATE Q

isopropyl2-((((((R)-1-(6-amino-9H-purin-9-yl)propan-2yl)oxy)methyl)(4-chlorophenoxy)-phosphoryl)amino)-2-methylpropanoate

A mixture of(R)-(((1-(6-amino-9H-purin-9-yl)propan-2-ypoxy)methyl)phosphonic acid(referred to herein as TFV, 19 g, 66.2 mmol), 4-chlorophenol (6.87 ml,69.5 mmol), INTERMEDIATE I (12.62 g, 69.5 mmol), and triethylamine (55.3ml, 397 mmol) in pyridine (221 mL) was heated to 60° C. for 5 minuteswith stirring. A freshly prepared solution of 2,2′-dipyridyl disulfide(Aldrithiol, 102 g, 463 mmol) and triphenylphosphine (121 g, 463 mmol)in pyridine (221 mL) was added to the mixture. The reaction was stirredat 60° C. over the weekend. The mixture was cooled to room temperatureand concentrated to remove most of the pyridine. The residue wasdirectly purified by gradient elution on silica gel (RediSep-Rf-1.5 kg,0-10% MeOH/DCM, 1 h gradient) to yield pure fractions which wereconcentrated to afford the title compound: LC/MS: [(M+1)]⁺=525.2.

INTERMEDIATE R

butyl2-((((((R)-1-(6-amino-9H-purin9-yl)propan-2-yl)oxy)methyl)(4-chlorophenoxy)phosphoryl)amino)-2-methylpropanoate

INTERMEDIATE R was prepared on a 3.13 mmol scale in a similar fashion tothat described for the synthesis of INTERMEDIATE Q starting from TFV,chlorophenol, and INTERMEDIATE J: LC/MS: [(M+1)]⁺=539.2.

INTERMEDIATE S

pentyl2-((((((R)-1-(6-amino-9H-purin-9-yl)propan-2-yl)oxy)methyl)(4-chlorophenoxy)phosphoryl)amino)-2-methylpropanoate

INTERMEDIATE S was prepared on a 25.7 mmol scale in a similar fashion tothat described for the synthesis of INTERMEDIATE Q starting from TFV,chlorophenol, and INTERMEDIATE K: LC/MS: [(M+1)]⁺=553.2.

INTERMEDIATE T

hexyl2-((((((R)-1-(6-amino-9H-purin-9yl)propan-2-yl)oxy)methyl)(4-chlorophenoxy)phosphoryl)amino)-2-methylpropanoate

INTERMEDIATE T was prepared on a 14.25 mmol scale in a similar fashionto that described for the synthesis of INTERMEDIATE Q starting from TFV,chlorophenol, and INTERMEDIATE L: LC/MS: [(MH+1)]⁺=567.3.

INTERMEDIATE U

isobutyl2-((((((R)-1-(6-amino-9H-purin-9-yl)propan-2yl)oxy)methyl)(4-chlorophenoxy)phosphoryl)amino)-2-methylpropanoate

INTERMEDIATE U was prepared on a 3.3 mmol scale in a similar fashion tothat described for the synthesis of INTERMEDIATE Q starting from TFV,chlorophenol, and INTERMEDIATE M LC/MS: [(M+1)]⁺=539.3.

INTERMEDIATE V

cyclobutyl2-((((((R)-1-(6-amino-9H-purin-9yl)propan-2-yl)oxy)methyl)(4-chlorophenoxy)phosphoryl)amino)-2-methylpropanopate

INTERMEDIATE V was prepared on a 0.42 mmol scale in a similar fashion tothat described for the synthesis of INTERMEDIATE Q starting from TFV,chlorophenol, and INTERMEDIATE N: LC/MS: [(MH+1)]⁺=537.2.

INTERMEDIATE W

cyclopentyl2-((((((R)-1-(6-amino-9H-purin-9-yl)propan-2-yl)oxy)methyl)(4-chlorophenoxy)phosphoryl)amino)-2-methylpropanoate

INTERMEDIATE W was preparedchlorophenoxv)nhosnhorvIlarnino)-2-methvInronarioate: INTERMEDIATE W wasprepared on a 0.13 mmol scale in a similar fashion to that described forthe synthesis of INTERMEDIATE Q starting from TFV, chtorophenol, andINTERMEDIATE O: LC/MS: [(M+1)]⁺=551.2.

INTERMEDIATE X

cyclohexyl2-((((((R)-1-(6-amino-9H-purin-9-yl)propan-2-yl)oxy)methyl)(4-chlorophenoxy)phosphoryl)amino)-2-methylpropanoate

INTERMEDIATE X was prepared on a 0.18 mmol scale in a similar fashion tothat described for the synthesis of INTERMEDIATE Q starting from TFV,chlorophenol, and INTERMEDIATE P: LC/MS: [(M+1)]⁺=565.2.

EXAMPLE 1A/B

-   isopropyl2-(((S)-((((R)-1-(6-amino-9H-purin-9-yl)propan-2-yl)oxy)methyl)(((R)-1-(cyclopentyloxy)-1-oxopropan-2-yl)amino)phosphoryl)amino)-2-methylpropanoate;    and-   isopropyl2-(((R)-((((R)-1-(6-amino-9H-purin-9-yl)propan-2-yl)oxy)methyl)(((R)-1-(cyclopentyloxy)-1-oxopropan-2-yl)amino)phosphoryl)amino)-2-methylpropanoate

isopropyl2-((((((R)-1-(6-amino-9H-purin-9-yl)propan-2-yl)oxy)methyl)(((R)-1-(cyclopentyloxy)-1-oxopropan-2-yl)amino)phosphoryl)amino)-2-methylpropanoate(1A/B):

A mixture of INTERMEDIATE Q (3.8 g, 7.24 mmol) and water (2.61 ml, 145mmol) in tetrahydrofuran (20 mL) was treated with DBU (1.64 mL, 10.86mmol) and stirred at room temperature for 2 hr. The mixture wasconcentrated and the residue was diluted with water (75 mL) and washedwith CHCl₃ (8×30 mL) to remove the chlorophenol byproduct. The desiredproduct stayed in the aqueous layer. The aqueous layer was concentrated,azeotroping twice with acetonitrile and once with toluene and was driedunder high vacuum to afford the unstable intermediate: LC/MS:[(M+1)]⁺=415.2. This intermediate was used immediately.

A mixture of the above intermediate (3 g, 7.24 mmol), INTERMEDIATE G(1.682 g, 8.69 mmol) and triethylamine (4.04 ml, 29.0 mmol) in pyridine(24 mL) was heated to 60° C. for 5 min. A freshly prepared solution of2,2′-dipyridyl disulfide (Aldrithiol, 4.78 g, 21.72 mmol) andtriphenylphosphine (5.70 g, 21.72 mmol) in pyridine (24 mL) was added tothe mixture. The reaction was stirred at 60° C. overnight. The mixturewas cooled to room temperature and concentrated to remove most of thepyridine. The crude material was purified directly by gradient elutionon silica gel (RediSep-Rf-120 g, 0-15% MeOH/CH₂Cl₂, 30 minute gradient)to afford the product (mixture of two isomers plus impurities) as afoam. The mixture was purified by reverse phase chromatography(Phenominex Gemini 5 um C18 50×250 mm column; 20-50% CH₃CN in a 5 mMsolution of NH₄HCO₃ over 30 min, 5 injections) and clean fractions werepartitioned between dichloromethane and water. The aqueous phases wereback-extracted once with dichloromethane. The combined organic phaseswere dried over Na₂SO₄, filtered and concentrated to afford Isomer 1A(faster eluting): ¹H NMR (500 MHz, DMSO): δ(ppm) 8.14 (d, J=9.8 Hz, 2H); 7.17 (s, 2 H); 5.06 (s, 1 H); 4.83-4.87 (m, 1 H); 4.15-4.27 (br m, 4H); 3.88 (m, 2 H); 3.61 (m, 1 H); 3.51 (m, 1 H); 1.80 (m, 2 H); 1.58 (m,6 H); 1.33 (d, J=4.0 Hz, 6 H); 1.22 (d, J=7.2 Hz, 3 H); 1.17 (dd, J=6.3,3.5 Hz, 6 H); 1.06 (d, J=6.2 Hz, 2 H); 0.88 (d, J=6.7 Hz, 1 H); ³¹P NMR(243 MHz, CD₃OD): δ(ppm) 22.16; LC/MS: [(M+1)]⁻=554.4; and Isomer 1B(slower eluting): ¹H NMR (500 MHz, DMSO): δ(ppm) 8.14 (d, 9.1 Hz, 2 H);7.17 (s, 2H); 5.07 (s, 1 H); 4.86 (t, J=6.3 Hz, 1 H); 4.36 (d, J=11.4Hz, 1 H); 4.26 (d, J=14.3 Hz, 1 H); 4.19 (t, J=6.7 Hz, 1 H); 4.10 (d,J=10.3 Hz, 1 H); 3.95 (s, 1 H); 3.86 (br s, 1 H); 3.57 (d, J=8.6 Hz, 2H); 1.81 (br s, 2 H); 1.55-1.63 (m, 6 H), 1.39 (s, 3 H); 1.35 (s, 3 H);1.16-1.19 (m, 9 H); 1.05 (d, J=6.2 Hz, 3 H); ⁻P NMR (243 MHz, CD₃OD):δ(ppm) 22.06; LC/MS: [(M+1)]⁺=554.4.

As noted above, absolute stereochemistry (R or 5) of each of theseparated stereoisomers was not determined. Therefore, A and B is notassigned to a particular IUPAC name.

EXAMPLE 2A/B

-   hexyl    2-(((S)-((((R)-1-(6-amino-9H-purin-9-yl)propan-2-yl)oxy)methyl)(((R)-1-isopropoxy-1-oxopropan-2-yl)amino)phosphoryl)amino)-2-methylpropanoate;    and-   hexyl    2-(((R)-((((R)-1-(6-amino-9H-purin-9-yl)propan-2-yl)oxy)methyl)(((R)-1-isopropoxy-1-oxopropan-2-yl)amino)phosphoryl)amino)-2-methylpropanoate

hexyl2-((((((R)-1-(6-amino-9H-purin-9-yl)propan-2-yl)oxy)methyl)(((R)-1-isopropoxy-1-oxopropan-2-yl)amino)phosphoryl)amino)-2-methylpropanoate(2A/B)

A mixture of INTERMEDIATE T (4 g, 7.05 mmol) and water (2.54 mL, 141mmol) in tetrahydrofuran (20 mL) was treated with DBU (1.75 ml, 11.64mmol) and stirred at room temperature for 2 hr. The mixture wasconcentrated and the residue was diluted with water (50 mL) and washedwith CH₂Cl₂ (2×100 mL) to remove chlorophenol byproduct. The desiredproduct stayed in the aqueous layer. The aqueous layer was concentratedazeotroping with MeCN (2×) and with toluene (1×), and was dried underhigh vacuum to afford the unstable intermediate: LC/MS: [(M+1)]⁺=457.3.This intermediate was used immediately.

A mixture of the above intermediate (3.22 g, 7.05 mmol), INTERMEDIATE A(2.37 g, 14.11 mmol) and triethylamine (3.93 mL, 28.2 mmol) in pyridine(23.5 mL) was heated to 60° C. for 5 min, A freshly prepared solution of2,2′-dipyridyl disulfide (Aldrithiol, 4.66 g, 21.16 mmol) andtriphenylphosphine (5.55 g, 21.16 mmol) in pyridine (23.5 mL) was addedto the mixture. The reaction was stirred at 60° C. overnight. Themixture was cooled to room temperature and concentrated to remove mostof the pyridine. The crude material was purified directly by gradientelution on silica gel (RediSep-Rf-330 g, 0-15% MeOH/CH₂Cl₂, 35 minutegradient) to afford the product (mixture of two isomers plus impurities)as a foam. (1.3 g) The mixture was purified by reverse phasechromatography (XBridge 10 um C18 30×250 mm column; 30-65% CH₃CN in a 5mM solution of NH₄HCO₃ over 30 min, 4 injections) and clean fractionswere partitioned between CH₂Cl₂ and water. The aqueous phases wereback-extracted once with CH₂Cl₂. The combined organic phases were driedover Na₂SO₄, filtered and concentrated to afford Isomer 2A (fastereluting): ¹H NMR (500 MHz, DMSO): δ (ppm) 8.14 (d, J=13.0 Hz, 2 H); 7.17(s, 2 H); 4.85-4.90 (m, 1 H); 4.13-4.29 (m, 4 H); 3.98-4.02 (m, 2 H);3.92 (d. J=7.4 Hz, 1 H); 3.83 (d, J=10.6 Hz, 1 H); 3.61 (dd, J=12.9, 8.3Hz, 1 H); 3.51 (dd, J=13.0, 8.8 Hz, 1 H); 1.51-1.57 (m, 2 H); 1.36 (s, 6H); 1.23 (d, J=8.0 Hz, 8 H); 1.17 (dd, J=6.4, 3.2 Hz, 7 H); 1.06 (d, J6.2 Hz, 3 H); 0.83 (d, J 7.4 Hz, 3 H); ³¹P NMR (243 MHz, CD₃OD): δ(ppm)22.23; LC/MS: [(M+1)]⁺=570.3; and Isomer 2B (slower eluting): ¹H NMR(500 MHz, DMSO): δ(ppm) 8.14 (d, J=10.0 Hz, 2H); 7.17 (s, 2 H);4.86-4.91 (m, 1 H); 4.35 (t, J=11.2 Hz, 1 H); 4.27 (dd, J=14.3, 4.0 Hz,1 H); 4.16-4.20 (m, 2 H); 4.02 (t, J=6.5 Hz, 2 H); 3.92-3.95 (m, 1 H);3.81-3.89 (m, 1 H); 3.57 (dd, J=8.6, 4.2 Hz, 2 H); 1.56 (p, J=7.0 Hz, 2H); 1.39 (d, J=15.2 Hz, 6 H); 1.25 (s, 6 H); 1.18-1.20 (m, 9 H); 1.05(d, J=6.2 Hz, 3 H); 0.84 (d, J=7.5 Hz, 3 H); ³¹P NMR (243 MHz, CD₃OD):δ(ppm) 22.07; LC/MS: [(M+1)]⁺=570.3. As noted above, absolutestereochemistry (R or S) of each of the separated stereoisomers was notdetermined. Therefore, A and B is not assigned to a particular IUPACname.

EXAMPLES 3-28

-   isopropyl    2-(((S)-((((R)-1-(6-amino-9H-purin-9-yl)propan-2-yl)oxy)methyl)(((R)-1-butoxy-1-oxopropan-2-yl)amino)phosphoryl)amino)-2-methylpropanoate;    and-   isopropyl    2-(((R)-((((R)-1-(6-amino-9H-purin-9-yl)propan-2-yl)oxy)methyl)(((R)-1-butoxy-1-oxopropan-2-yl)amino)phosphoryl)amino)-2-methylpropanoate    (3A/B)

isopropyl2-((((((R)-1-(6-amino-9H-purin-9-yl)propan-2-yl)oxy)methyl)(((R)-1-butoxy-1-oxopropan-2-yl)amino)phosphoryl)amino)-2-methylpropanoate(3A/B);

A mixture of INTERMEDIATE Q (10.3 g, 19.6 mmol) and water (7.07 ml, 392mmol) in tetrahydrofuran (49 mL) was treated with DBU (3.11 mL, 20.6mmol) and stirred at room temperature for 3 hr. The mixture wasconcentrated and the residue was diluted with water (300 mL) and washedwith CHCl₃ (8×500 mL) to remove chlorophenol byproduct. The desiredproduct stayed in the aqueous layer. The aqueous layer was concentratedazeotroping with acetonitrile twice and once with toluene and was driedunder high vacuum to afford the unstable intermediate: LC/MS:[(M+1)]⁺=415.2. This intermediate was used immediately.

A mixture of the above intermediate (8,1 g, 19.5 mmol), INTERMEDIATE D(4.1 g, 22.5 mmol) and triethylamine (10.9 mL, 78.0 mmol) in pyridine(65 mL) was heated to 60° C. for 5 min. A freshly prepared solution of2,2′-dipyridyl disulfide (Aldrithiol, 12.9 g, 58.6 mmol) andtriphenylphosphine (15.4 g, 58.6 mmol) in pyridine (65 mL) was added tothe mixture. The reaction was stirred at 60° C. overnight. The mixturewas cooled to room temperature and concentrated to remove most of thepyridine. The crude material was purified directly by gradient elutionon silica gel (RediSep-Rf-330 g, 0-15% MeOH/CH₂Cl₂, 30-min gradient) toafford the product (mixture of two isomers plus impurities) as a foam.The mixture was purified by reverse phase chromatography (PhenominexGemini 5 um C18 50×250 mm column; 20-50% CH₃CN in a 5 mM solution ofNH₄HCO₃ over 30 min, 5 injections) and clean fractions were partitionedbetween CH₂Cl₂ and water. The aqueous phases were back-extracted oncewith CH₂Cl₂. The combined organic phases were dried over Na₂SO₄,filtered and concentrated to afford Isomer 3A (faster eluting): ¹H NMR(500 MHz, DMSO): δ (ppm) 8.14 (d,=10.5 Hz, 2 H); 7.17 (s, 2 H);4.81-4.86 (m, 1 H); 4.26 (t, J=11.4 Hz, 2 H); 4.14-4.18 (m, 2 H);3.98-4.07 (m, 2 H); 3.91 (m, 1 H); 3.61 (dd, J=12.9, 8.4 Hz, 1 H); 3.51(dd, J=12.8, 8.8 Hz, 2 H); 1.54 (p, J=7.1 Hz, 2 H); 1.29-1.34 (m, 8 H);1.25 (d, J=7.1 Hz, 3 H), 1.17 (dd, J=6.3, 2.9 Hz, 6 H); 1.06 (d, J=6.2Hz, 3 H); 0.87 (t, J=7.4 Hz, 3 H); ³¹P NMR (243 MHz, CD₃OD): δ(ppm)22.16; LC/MS: [(M+1)]⁺=542.3; and isomer 3B (slower eluting): ¹H NMR(500 MHz, DMSO): δ(ppm) 8.14 (d, J=9.4 Hz, 2 H); 7.17 (s, 2 H);4.83-4.88 (m, 1 H); 4.40 (t, J=11.3 Hz, 1 H); 4.26 (dd, J=14.3, 4.0 Hz,1 H); 4.18 (dd, J=14.5, 5.9 Hz, 1 H); 4.13 (d, J=10.3 Hz, 1 H);4.02-4.07 (m, 2 H); 3.88-3.96 (m, 2 H); 3.55-3.58 (m, 2 H); 1.52-1.58(m, 2 H); 1.39 (s, 3 H); 1.35 (s, 3 H); 1.30-1.33 (m, 3 H); 1.16-1.21(m, 8 H); 1.05 (d, J=6.2 Hz, 3 H); 0.88 (t, J=7.4 Hz, 3 H); ³¹P NMR (243MHz, CD₃OD): -δ(ppm) 22.04; LC/MS: [(M+1)]⁺=542.3. As noted above,absolute stereochemistry (R or S) of each of the separated stereoisomerswas not determined. Therefore, A and B is not assigned to a particularIUPAC name.

The compounds in Table 1 were prepared in an analogous fashion to thatdescribed for the examples above. The isomers were separated bypreparative HPLC or/and preparative chiral SFC.

TABLE 1 Compound Structure Name LC/MS (M + 1)⁺  4A/B

isopropyl 2-(((S)-(((((R)-1- (6-amino-9H-purin-9- yl)propan-2-yl)oxy)methyl)(((R)-1- isopropoxy-1-oxopropan-2-yl)amino)phosphoryl)amino)- 2-methylpropanoate and isopropyl2-(((R)-((((R)-1- (6-amino-9H-purin-9- yl)propan-2-yl)oxy)methyl)(((R)-1 isopropoxy-1-oxopropan-2-yl)amino)phosphoryl)amino)- 2-methylpropanoate 528.3 and 528.3  5A/B

isopropyl 2-(((S)-((((R)-1- (6-amino-9H-purin-9- yl)propan-2-yl)oxy)methyl)(((R)-1- isobutoxy-1-oxopropan-2-yl)amino)phosphoryl)amino)- 2-methylpropanoate and isopropyl2-(((R)-((((R)-1- (6-amino-9H-purin-9- yl)propan-2-yl)oxy)methyl)(((R)-1- isobutoxy-1-oxopropan-2-yl)amino)phosphoryl)amino)- 2-methylpropanoate 542.3 and 542.3  6A/B

isopropyl 2-(((S)-((((R)-1- (6-amino-9H-purin-9- yl)propan-2-yl)oxy)methyl)(((R)-1- (cyclohexyloxy)-1- oxopropan-2-yl)amino)phosphoryl)amino)- 2-methylpropanoate and isopropyl2-(((R)-((((R)-1- (6-amino-9H-purin-9- yl)propan-2-yl)oxy)methyl)(((R)-1- (cyclohexyloxy)-1- oxopropan-2-yl)amino)phosphoryl)amino)- 2-methylpropanoate 568.3 and 568.3  7A/B

isopropyl 2-(((S)-((((R)-1- (6-amino-9H-purin-9- yl)propan-2-yl)oxy)methyl)(((R)-1- (hexyloxy)-1-oxopropan-2-yl)amino)phosphoryl)amino)- 2-methylpropanoate and isopropyl2-(((R)-((((R)-1- (6-amino-9H-purin-9- yl)propan-2-yl)oxy)methyl)(((R)-1- (hexyloxy)-1-oxopropan-2-yl)amino)phosphoryl)amino)- 2-methylpropanoate 570.3 and 570.4  8A/B

isopropyl 2-(((S)-((((R)-1- (6-amino-9H-purin-9- yl)propan-2-yl)oxy)methyl)(((R)-1- oxo-1-(pentyloxy)propan-2-yl)amino)phosphoryl)amino)- 2-methylpropanoate and isopropyl2-(((R)-((((R)-1- (6-amino-9H-purin-9- yl)propan-2-yl)oxy)methyl)(((R)-1- oxo-1-(pentyloxy)propan-2-yl)amino)phosphoryl)amino)- 2-methylpropanoate 556.4 and 556.4  9A/B

pentyl 2-(((S)-((((R)-1-(6- amino-9H-purin-9- yl)propan-2-yl)oxy)methyl)(((R)-1- butoxy-1-oxopropan-2- yl)amino)phosphoryl)amino)-2-methylpropanoate and pentyl 2-(((R)-((((R)-1-(6- amino-9H-purin-9-yl)propan-2- yl)oxy)methyl)(((R)-1- butoxy-1-oxopropan-2-yl)amino)phosphoryl)amino)- 2-methylpropanoate 570.3 and 570.3 10A/B

hexyl 2-(((S)-((((R)-1-(6- amino-9H-purin-9- yl)propan-2-yl)oxy)methyl)(((R)-1- isobutoxy-1-oxopropan-2-yl)amino)phosphoryl)amino)- 2-methylpropanoate and hexyl2-(((R)-((((R)-1-(6- amino-9H-purin-9- yl)propan-2-yl)oxy)methyl)(((R)-1- isobutoxy-1-oxopropan-2-yl)amino)phosphoryl)amino)- 2-methylpropanoate 584.4 and 584.4 11A/B

hexyl 2-(((S)-((((R)-1-(6- amino-9H-purin-9- yl)propan-2-yl)oxy)methyl)(((R)-1- butoxy-1-oxopropan-2- yl)amino)phosphoryl)amino)-2-methylpropanoate and hexyl 2-(((R)-((((R)-1-(6- amino-9H-purin-9-yl)propan-2- yl)oxy)methyl)(((R)-1- butoxy-1-oxopropan-2-yl)amino)phosphoryl)amino)- 2-methylpropanoate 584.4 and 584.4 12A/B

hexyl 2-(((S)-((((R)-1-(6- amino-9H-purin-9- yl)propan-2-yl)oxy)methyl)(((R)-1- oxo-1-(pentyloxy)propan-2-yl)amino)phosphoryl)amino)- 2-methylpropanoate and hexyl2-(((R)-((((R)-1-(6- amino-9H-purin-9- yl)propan-2-yl)oxy)methyl)(((R)-1- oxo-1-(pentyloxy)propan-2-yl)amino)phosphoryl)amino)- 2-methylpropanoate 598.4 and 598.4 13A/B

hexyl 2-(((S)-((((R)-1-(6- amino-9H-purin-9- yl)propan-2-yl)oxy)methyl)(((R)-1- (hexyloxy)-1-oxopropan-2-yl)amino)phosphoryl)amino)- 2-methylpropanoate and hexyl2-(((R)-((((R)-1-(6- amino-9H-purin-9- yl)propan-2-yl)oxy)methyl)(((R)-1- (hexyloxy)-1-oxopropan-2-yl)amino)phosphoryl)amino)- 2-methylpropanoate 612.4 and 612.4 14A/B

hexyl 2-(((S)-((((R)-1-(6- amino-9H-purin-9- yl)propan-2-yl)oxy)methyl)(((R)-1- (cyclopentyloxy)-1- oxopropan-2-yl)amino)phosphoryl)amino)- 2-methylpropanoate and hexyl2-(((R)-((((R)-1-(6- amino-9H-purin-9- yl)propan-2-yl)oxy)methyl)(((R)-1- (cyclopentyloxy)-1- oxopropan-2- 596.4 and 596.4yl)amino)phosphoryl)amino)- 2-methylpropanoate 15A/B

hexyl 2-(((S)-((((R)-1-(6- amino-9H-purin-9- yl)propan-2-yl)oxy)methyl)(((R)-1- (cyclohexyloxy)-1- oxopropan-2-yl)amino)phosphoryl)amino)- 2-methylpropanoate and hexyl2-(((R)-((((R)-1-(6- amino-9H-purin-9- yl)propan-2-yl)oxy)methyl)(((R)-1- (cyclohexyloxy)-1- oxopropan-2-yl)amino)phosphoryl)amino)- 610.4 and 610.4 2-methylpropanoate 16A/B

pentyl 2-(((S)-((((R)-1-(6- amino-9H-purin-9- yl)propan-2-yl)oxy)methyl)(((R)-1- oxo-1-(pentyloxy)propan-2-yl)amino)phosphoryl)amino)- 2-methylpropanoate and pentyl2-(((R)-((((R)-1-(6- amino-9H-purin-9- yl)propan-2-yl)oxy)methyl)(((R)-1- oxo-1-(pentyloxy)propan-2-yl)amino)phosphoryl)amino)- 2-methylpropanoate 584.4 and 584.4 17A/B

pentyl 2-(((S)-((((R)-1-(6- amino-9H-purin-9- yl)propan-2-yl)oxy)methyl)(((R)-1- isopropoxy-1-oxopropan-2-yl)amino)phosphoryl)amino)- 2-methylpropanoate and pentyl2-(((R)-((((R)-1-(6- amino-9H-purin-9- yl)propan-2-yl)oxy)methyl)(((R)-1- isopropoxy-1-oxopropan-2-yl)amino)phosphoryl)amino)- 2-methylpropanoate 556.3 and 556.3 18A/B

isopropyl 2-(((S)-((((R)-1- (6-amino-9H-purin-9- yl)propan-2-yl)oxy)methyl)(((R)-1- cyclobutoxy-1-oxopropan-2-yl)amino)phosphoryl)amino)- 2-methylpropanoate and isopropyl2-(((R)-((((R)-1- (6-amino-9H-purin-9- yl)propan-2-yl)oxy)methyl)(((R)-1- cyclobutoxy-1-oxopropan-2-yl)amino)phosphoryl)amino)- 2-methylpropanoate 540.2 and 540.2 19A/B

pentyl 2-(((S)-((((R)-1-(6- amino-9H-purin-9- yl)propan-2-yl)oxy)methyl)(((R)-1- cyclobutoxy-1-oxopropan-2-yl)amino)phosphoryl)amino)- 2-methylpropanoate and pentyl2-(((R)-((((R)-1-(6- amino-9H-purin-9- yl)propan-2-yl)oxy)methyl)(((R)-1- cyclobutoxy-1-oxopropan-2-yl)amino)phosphoryl)amino)- 2-methylpropanoate 568.4 and 568.4 20A/B

isobutyl 2-(((S)-((((R)-1- (6-amino-9H-purin-9- yl)propan-2-yl)oxy)methyl)(((R)-1- cyclobutoxy-1-oxopropan-2-yl)amino)phosphoryl)amino)- 2-methylpropanoate and isobutyl2-(((R)-((((R)-1- (6-amino-9H-purin-9- yl)propan-2-yl)oxy)methyl)(((R)-1- cyclobutoxy-1-oxopropan-2-yl)amino)phosphoryl)amino)- 2-methylpropanoate 554.2 and 554.2 21A/B

hexyl 2-(((S)-((((R)-1-(6- amino-9H-purin-9- yl)propan-2-yl)oxy)methyl)(((R)-1- cyclobutoxy-1-oxopropan-2-yl)amino)phosphoryl)amino)- 2-methylpropanoate and hexyl2-(((R)-((((R)-1-(6- amino-9H-purin-9- yl)propan-2-yl)oxy)methyl)(((R)-1- cyclobutoxy-1-oxopropan-2-yl)amino)phosphoryl)amino)- 2-methylpropanoate 582.2 and 582.2 22A/B

propyl 2-(((S)-((((R)-1-(6- amino-9H-purin-9- yl)propan-2-yl)oxy)methyl)(((R)-1- cyclobutoxy-1-oxopropan-2-yl)amino)phosphoryl)amino)- 2-methylpropanoate and propyl2-(((R)-((((R)-1-(6- amino-9H-purin-9- yl)propan-2-yl)oxy)methyl)(((R)-1- cyclobutoxy-1-oxopropan-2-yl)amino)phosphoryl)amino)- 2-methylpropanoate 540.2 and 540.2 23A/B

butyl 2-(((S)-((((R)-1-(6- amino-9H-purin-9- yl)propan-2-yl)oxy)methyl)(((R)-1- cyclobutoxy-1-oxopropan-2-yl)amino)phosphoryl)amino)- 2-methylpropanoate and butyl2-(((R)-((((R)-1-(6- amino-9H-purin-9- yl)propan-2-yl)oxy)methyl)(((R)-1- cyclobutoxy-1-oxopropan-2-yl)amino)phosphoryl)amino)- 2-methylpropanoate 554.2 and 554.2 24A/B

pentyl 2-(((S)-((((R)-1-(6- amino-9H-purin-9- yl)propan-2-yl)oxy)methyl)(((R)-1- (cyclopentyloxy)-1- oxopropan-2-yl)amino)phosphoryl)amino)- 2-methylpropanoate and pentyl2-(((R)-((((R)-1-(6- amino-9H-purin-9- yl)propan-2-yl)oxy)methyl)(((R)-1- (cyclopentyloxy)-1- oxopropan-2-yl)amino)phosphoryl)amino)- 2-methylpropanoate 582.4 and 582.4 25A/B

pentyl 2-(((S)-((((R)-1-6- amino-9H-purin-9- yl)propan-2-yl)oxy)methyl)(methyl((R)- 1-oxo-1- (pentyloxy)propan-2-yl)amino)phosphoryl)amino)- 2-methylpropanoate and pentyl2-(((R)-((((R)-1-(6- amino-9H-purin-9- yl)propan-2-yl)oxy)methyl)(methyl((R)- 1-oxo-1- (pentyloxy)propan-2-yl)amino)phosphoryl)amino)- 598.4 and 598.4 2-methylpropanoate 26A/B

pentyl (((S)-(((R)-1-(6- amino-9H-purin-9- yl)propan-2-yl)oxy)methyl)((2-methyl- 1-oxo-1- (pentyloxy)propan-2-yl)amino)phosphoryl)-D- prolinate and pentyl (((R)-(((R)-1-(6-amino-9H-purin-9- yl)propan-2- yl)oxy)methyl)((2-methyl- 1-oxo-1-(pentyloxy)propan-2- yl)amino)phosphoryl)-D- prolinate 610.4 and 610.427A/B

cyclohexyl 2-(((S)-((((R)- 1-(6-amino-9H-purin-9- yl)propan-2-yl)oxy)methyl)(((R)-1- (cyclopentyloxy)-1- oxopropan-2-yl)amino)phosphoryl)amino)- 2-methylpropanoate and cyclohexyl2-(((R)-((((R)- 1-(6-amino-9H-purin-9- yl)propan-2-yl)oxy)methyl)(((R)-1- (cyclopentyloxy)-1- oxopropan-2-yl)amino)phosphoryl)amino)- 2-methylpropanoate 594.4 and 594.3 28A/B

cyclobutyl 2-(((S)-((((R)- 1-(6-amino-9H-purin-9- yl)propan-2-yl)oxy)methyl)(((R)-1- (cyclopentyloxy)-1- oxopropan-2-yl)amino)phosphoryl)amino)- 2-methylpropanoate and cyclobutyl2-(((R)-((((R)- 1-(6-amino-9H-purin-9- yl)propan-2-yl)oxy)methyl)(((R)-1- (cyclopentyloxy)-1- oxopropan-2-yl)amino)phosphoryl)amino)- 566.3 and 566.3 2-methylpropanoate

As noted above, absolute stereochemistry (R or S) of each of theseparated stereoisomers was not determined. Therefore, A and B is notassigned to a particular IUPAC name in this table.

EXAMPLE 29 Assessing Antiviral Potency in a Multiple Round HIV-1Infection Assay (Viking Assay)

The antiviral activity of the tenofovir prodrugs of the Examples hereinwas assessed in an assay that measures the rate of replication of HIV incell culture, termed the Viking assay (VIral KINetics in Green cells)and performed as follows. HIV-1 replication was monitored usingMT4-gag-GFP clone D3 (hereafter designated MT4-GFP; GFP is greenfluorescent protein), which are MT-4 cells modified to harbor a GFPreporter gene, the expression of which is dependent on the HIV-1expressed proteins tat and rev; see Wang Y-J et al., 2010J VirologicalMethods 165:230-237. Productive infection of an MT4-GFP cell with HIV-1results in GFP expression approximately 24 h post-infection. MT4-GFPcells were maintained at 37° C./5% CO₂/90% relative humidity in RPMI1640 supplemented with 10% fetal bovine serum, 100 U/mlpenicillin/streptomycin, and 400 μg/ml G418 to maintain the reportergene. For infections, MT4-GFP cells were placed in the same mediumlacking G418 and infected overnight with HIV-1 (H9/IIIB strain) virus atan approximate multiplicity of infection of 0.01 in the same incubationconditions. Cells were then washed and re-suspended in either RPMI 1640supplemented with 10% or 50% normal human serum (NHS) at 1.6×10⁵cells/mL (10% NHS or 50% NHS, respectively). Compound plates wereprepared by dispensing compounds dissolved in DMSO into wells of 384well poly-D-lysine-coated plates (0.2 μl/well) using an ECHO acousticdispenser. Each compound was tested in a 10-point serial 3-fold dilution(typical final concentrations: 8.4 μM-0.42 nM). Controls included noinhibitor (DMSO only) and a combination of three antiviral agents(efavirenz, indinavir, and an integrase strand transfer inhibitor atfinal concentrations of 4 μM each). Cells were added (50 μL/well) tocompound plates and the infected cells were maintained at 37′C/5%CO2/90% relative humidity.

Infected cells were quantified at two time points, ˜48 h and ˜72 hpost-infection, by counting the number of green cells in each well usingan Acumen eX3 scanner. The increase in the number of green cells over˜24 h period gives the reproductive ratio, R0, which is typically 5-15and has been shown experimentally to be in logarithmic phase (data notshown). R0 is calculated for each well, and IC₅₀s determined bynon-linear 4-parameter curve fitting. Assay IC₅₀ results are shown inTable 2.

EXAMPLE 30 Prodrug Stability Assay in Bio-relevant Media

The following assay was employed to evaluate the stability of theprodrugs in simulated gastrointestinal tract conditions. Preparation offasted state simulated intestinal fluid (FaSSIF) using Phares SIF Powderwas carried out according to protocols from Phare Drug Delivery AG(Baselland, Switzerland). For sample preparation, 10 μL stock solutions(10 mM) of prodrug substance in DMSO was added to 990 μL of 0.5 mg/mLPancreatin solution (Fisher CAS#8049-47-6) in FaSSIF. Two samples wereprepared for each compound. If the sample was a clear solution, it wasdirectly analyzed by HPLC. If the sample was not clear, the sample wasdiluted with 100% ACN, maintained at 37° C. and observed 5 h later. Ifthe sample was clear, HPLC analysis was directly performed. If thesample was still not clear, the sample was diluted with 100% ACN andassayed by HPLC. All the samples were vortexed for 3 min and observedbefore injection. For the diluted samples, the area is multiplied by afactor when the data is analyzed. The analysis was carried out with anAgilent 1100 series HPLC with autosampler. The column was usually aPoroshell 120 EC-C18, 4.6×50 mm, 2.7 μm. The flow rate was 1.8 mL/min,and the injection volume was 5 or 10 μL. UV detection was in the range210-400 nm. The mobile phase consisted of solvent A (water plus 10 mMtetrabutylammonium bromide) and solvent B (acetonitrile) with a gradientof 90% solvent A at 0 min, changing to 95% solvent B over 6 min,maintaining for 1.5 min, then reverting to 90% solvent A over 1.6 min.The HPLC peak area of the prodrug at 5 h was divided by the HPLC peakarea of the prodrug at 0 h, to generate the % claimed parent ratio,which is summarized in Table 2. for GI Tract stability.

EXAMPLE 31 Pharmacokinetic Studies in Dogs—In Vivo Dog PK

Prodrugs were administered to beagle dogs through intravenous (IV) andoral (P.O.) administrations in a non-crossover manner. The IV dose wasprepared in 20% hydroxypropyl β-cyclodextrin (HPBCD) and wasadministered at 1 mg/kg QD via the cephalic or saphenous vein. The P.O.dose was prepared in 10% polysorbate 80 (Tween 80) and was administeredat 10 mg/kg QD via gavage.

Blood samples were serially collected following dose administration forup to 48 hr and plasma was separated by centrifugation. Theconcentrations of prodrugs in dog plasma were determined by a LC-MS/MSassay following a protein precipitation step and addition of anappropriate internal standard (labetalol, imipramine or diclofenac).Quantification was done by determining peak area-ratios of the prodrugsand tenofovir to the internal standard. Additional blood sample(s) wascollected following dose administration for up to 24 hr. Peripheralblood mononuclear cells (PRNICs) were isolated by centrifugation, usingtubes and reagents specified for the application. The concentrations oftenofovir and/or its phosphate conjugate(s) in PBMCs were determined byan LC-MS/MS assay following a protein precipitation step and addition ofan appropriate internal standard (labetalol, imipramine or diclofenac).Quantification was done by determining peak area-ratios of tenofovirand/or its phosphate conjugate(s) to the internal standard.

Pharmacokinetic parameters were obtained using non-compartmental methods(Watson®). The area under the plasma concentration-time curve(AUC_(0-t)) was calculated from the first time point (0 min) up to thelast time point with measurable drug concentration using the lineartrapezoidal or linear/log-linear trapezoidal rule. The IV plasmaclearance was calculated by dividing the dose by AUC_(0-inf). Theterminal half-life of elimination was determined by un-weighted linearregression analysis of the log-transformed data. The time points fordetermination of half-life were selected by visual inspection of thedata. The volume of distribution at steady state (Vd_(ss)) was obtainedfrom the product of plasma clearance and mean residence time (determinedby dividing the area under the first moment curve by the area under thecurve). The maximum plasma concentration (C_(max)) and the time at whichmaximum concentration occurred (T_(max)) were obtained by inspection ofthe plasma concentration-time data. Absolute oral bioavailability (% F)was determined from dose-adjusted IV and P.O. AUC ratios of the prodrug.

Without wishing to be bound by theory, it is believed that the compoundsof

Formula I are converted by one or more mechanisms to the correspondingmonophosphate or diphosphate of tenofovir. In vivo, tenofovir (TFV) isphosphorylated intracellularly to its active anabolite, tenofovirdiphosphate (TFV-DP), which suppresses viral replication via inhibitingthe activity of HIV reverse transcriptase to impart antiviral efficacy.Table 2 shows in vivo dog PK data in the form of TFV-DP concentrations(μM) in dog PRMCs at 24 h following a 10 mg/kg P.O. dose of theindicated prodrug.

TABLE 2 Viking, IC50 Viking, IC50 (50% NHS) GI Tract In Vivo Dog Example(10% NHS) (nM) (nM) stability (%) PK (μM)  1A 19.3 44.3  1B 2.4 6.2 95.924.5  2A 15.9 78 100 31.0  2B 1.3 5.2 94.2 18.2  3A 20.7 71.4  3B 3.820.5 95.6 15.3  4A 417.8 1396  4B 74.6 122.3  5A 33.1 129.4  5B 2.5 6.410.7  6A 100.6 500.5  6B 1.9 5.6 11.7  7A 10.4 57.8  7B 4.7 28.8  8A12.6 41.4  8B 3.9 20.7  9A 6.1 19.8 100  9B 1.0 4.2 73.9 10A 6.6 72.510B 0.7 2.0 66.8 11A 9.9 99.0 11B 0.8 2.6 12A 7.3 54.4 12B 0.5 3.3 13A7.2 40.4 99.1 13B 83.8 14A 9.8 67.2 14B 0.6 2.0 47.2 15A 7.7 25.9 15B0.7 2.1 51.2 16A 10.0 60.1 16B 0.9 4.2 97.8 17A 15.9 78.0 100 17B 1.35.2 94.2 18A 20.0 34.2 18B 11.0 25.8 97.4 26.5 19A 6.9 27.4 19B 2.5 10.849.3 20A 5.9 17.4 98.1 20B 1.7 5.9 71.0 21A 4.4 14.9 21B 0.7 2.8 22A11.6 50.9 98.2 18.8 22B 4.4 15.4 72.6 16.0 23A 6.7 35.5 23B 6.0 13.658.6 24A 3.2 30.6 97.8 24B 0.9 3.9 82.7 13.0 25A 25B 3.6 9.6 94.4 26A5.3 50.3 26B 4.4 19.5 92.5 27A 12.9 64.5 27B 1.6 4.7 93.5 28A 5.4 16.128B 2.7 3.3 90.2

What is claimed is:
 1. A compound of structural Formula I:

or pharmaceutically acceptable salt thereof, wherein: R^(A) is aD-proline ester residue of formula (ii):

R³ is (a) —C₁₋₁₀alkyl unsubstituted or substituted with one to threesubstituents independently selected from fluoro, chloro, bromo, —OR⁷,—SH, —NR⁸R⁹, —C₃₋₆cycloalkyl or spiro—C₃₋₆cycloalkyl, (b) —CH₂-phenylunsubstituted or substituted with one to three substituentsindependently selected from fluoro, chloro, bromo, —OR¹⁰, —SH, —NR⁸R⁹ or—C₁₋₃alkyl, (c) —C₃₋₈cycloalkyl unsubstituted or substituted with one tothree substituents independently selected from fluoro, chloro, bromo,—OR¹⁰, —SH, —NR⁸R⁹ or —C₁₋₃alkyl, (d) aryl unsubstituted or substitutedwith one to three substituents independently selected from fluoro,chloro, bromo, —OR¹⁰, —SH, —NR⁸R⁹ or —C₁₋₃alkyl, (e)—C₁₋₅alkyl-X—C₁₋₅alkyl wherein X is O, S or NH, (f) heteroarylunsubstituted or substituted with one to three substituentsindependently selected from fluoro, chloro, bromo, —OR¹⁰, —SH, —NR⁸R⁹ or—C₁₋₃alkyl, or (g) a heterocyclic ring unsubstituted or substituted withone to three substituents independently selected from fluoro, chloro,bromo, —OR¹⁰, —SH, —NR⁸R⁹ or —C₁₋₃alkyl; R^(B) is a geminallydi-substituted amino acid ester residue of formula (iii):

R⁴ and R⁵ are each independently selected from (a) —C₁₋₄alkyl, (b)—C₁₋₄alkyl substituted with —OH, —SH, —SCH₃, —NH₂, —NH—C(═NH)—NH₂, (c)—CH₂-phenyl, (d) —CH₂-phenol, (e) —(CH₂)₁₋₂—COOH, (f) —(CH₂)₁₋₂—CONH₂,(g) —CH₂-1H-indole, (h) —CH₂-imidazole, (i) aryl, or (j) heteroaryl; orR⁴ and R⁵ are joined together with the carbon to which they are bothattached to form —C₃₋₆cycloalkyl or a 4 to 6-membered heterocyclic ring;R⁶ is (a) —C₁₋₁₀alkyl unsubstituted or substituted with one to threesubstituents independently selected from fluoro, chloro, bromo, —OR¹¹,—SH, —NR¹²R¹³, —C₃₋₆cycloalkyl or spiro-C₃₋₆cycloalkyl, (b) —CH₂-phenylunsubstituted or substituted with one to three substituentsindependently selected from fluoro, chloro, bromo, —OR¹⁴, —SH, —NR¹²R¹³,—C₁₋₃alkyl, (c) —C₃₋₈cycloalkyl unsubstituted or substituted with one tothree substituents independently selected from fluoro, chloro, bromo,—OR¹⁴, —SH, —NR¹²R¹³ or —C₁₋₃alkyl, (d) aryl unsubstituted orsubstituted with one to three substituents independently selected fromfluoro, chloro, bromo, —OR¹⁴, —SH, —NR¹²R¹³ or —C₁₋₃alkyl, (e)—C₁₋₅alkyl-X—C₁₋₅alkyl wherein X is O, S or NH; (f) heteroarylunsubstituted or substituted with one to three substituentsindependently selected from fluoro, chloro, bromo, —OR¹⁴, —SH, —NR¹²R¹³or —C₁₋₃alkyl, or (g) a heterocyclic ring unsubstituted or substitutedwith one to three substituents independently selected from fluoro,chloro, bromo, —OR¹⁴, —SH, —NR¹²R¹³ or —C₁₋₃alkyl; R⁷ and R¹¹ are eachindependently selected from —H or —C₃₋₆cycloalkyl; R⁸ and R⁹ are eachindependently selected from —H, —C₁₋₃alkyl or —C₃₋₆cycloalkyl; R¹² andR¹³ are each independently selected from —H, —C₁₋₃alkyl or—C₃₋₆cycloalkyl; and R¹⁰ and R¹⁴ are each independently selected from—H, —C₁₋₃alkyl or —C₃₋₆cycloalkyl.
 2. A compound which is selected fromthe following:

or a pharmaceutically acceptable salt thereof.
 3. A compound which isselected from the following:isopropyl2-(((S)-((((R)-1-(6-amino-9H-purin-9-yl)propan-2-yl)oxy)methyl)(((R)-1-(cyclopentyloxy)-1-oxopropan-2-yl)amino)phosphoryl)amino)-2-methylpropanoate;isopropyl2-(((R)-((((R)-1-(6-amino-9H-purin-9-yl)propan-2-yl)oxy)methyl)(((R)-1-(cyclopentyloxy)-1-oxopropan-2-yl)amino)phosphoryl)amino)-2-methylpropanoate;hexyl2-(((S)-((((R)-1-(6-amino-9H-purin-9-yl)propan-2-yl)oxy)methyl)(((R)-1-isopropoxy-1-oxopropan-2-yl)amino)phosphoryl)amino)-2-methylpropanoate;hexyl2-(((R)-((((R)-1-(6-amino-9H-purin-9-yl)propan-2-yl)oxy)methyl)(((R)-1-isopropoxy-1-oxopropan-2-yl)amino)phosphoryl)amino)-2-methylpropanoate;isopropyl2-(((S)-((((R)-1-(6-amino-9H-purin-9-yl)propan-2-yl)oxy)methyl)(((R)-1-butoxy-1-oxopropan-2-yl)amino)phosphoryl)amino)-2-methylpropanoate;isopropyl2-(((R)-((((R)-1-(6-amino-9H-purin-9-yl)propan-2-yl)oxy)methyl)(((R)-1-butoxy-1-oxopropan-2-yl)amino)phosphoryl)amino)-2-methylpropanoate;isopropyl2-(((S)-((((R)-1-(6-amino-9H-purin-9-yl)propan-2-yl)oxy)methyl)(((R)-1-isopropoxy-1-oxopropan-2-yl)amino)phosphoryl)amino)-2-methylpropanoate;isopropyl2-(((R)-((((R)-1-(6-amino-9H-purin-9-yl)propan-2-yl)oxy)methyl)(((R)-1-isopropoxy-1-oxopropan-2-yl)amino)phosphoryl)amino)-2-methylpropanoate;isopropyl2-(((S)-((((R)-1-(6-amino-9H-purin-9-yl)propan-2-yl)oxy)methyl)(((R)-1-isobutoxy-1-oxopropan-2-yl)amino)phosphoryl)amino)-2-methylpropanoate;isopropyl2-(((R)-((((R)-1-(6-amino-9H-purin-9-yl)propan-2-yl)oxy)methyl)(((R)-1-isobutoxy-1-oxopropan-2-yl)amino)phosphoryl)amino)-2-methylpropanoate;isopropyl2-(((S)-((((R)-1-(6-amino-9H-purin-9-yl)propan-2-yl)oxy)methyl)(((R)-1-(cyclohexyloxy)-1-oxopropan-2-yl)amino)phosphoryl)amino)-2-methylpropanoate;isopropyl2-(((R)-((((R)-1-(6-amino-9H-purin-9-yl)propan-2-yl)oxy)methyl)(((R)-1-(cyclohexyloxy)-1-oxopropan-2-yl)amino)phosphoryl)amino)-2-methylpropanoate;isopropyl2-(((S)-((((R)-1-(6-amino-9H-purin-9-yl)propan-2-yl)oxy)methyl)(((R)-1-(hexyloxy)-1-oxopropan-2-yl)amino)phosphoryl)amino)-2-methylpropanoate;isopropyl2-(((R)-((((R)-1-(6-amino-9H-purin-9-yl)propan-2-yl)oxy)methyl)(((R)-1-(hexyloxy)-1-oxopropan-2-yl)amino)phosphoryl)amino)-2-methylpropanoate;isopropyl2-(((S)-((((R)-1-(6-amino-9H-purin-9-yl)propan-2-yl)oxy)methyl)(((R)-1-oxo-1-(pentyloxy)propan-2-yl)amino)phosphoryl)amino)-2-methylpropanoate;isopropyl2-(((R)-((((R)-1-(6-amino-9H-purin-9-yl)propan-2-yl)oxy)methyl)(((R)-1-oxo-1-(pentyloxy)propan-2-yl)amino)phosphoryl)amino)-2-methylpropanoate;pentyl2-(((S)-((((R)-1-(6-amino-9H-purin-9-yl)propan-2-yl)oxy)methyl)(((R)-1-butoxy-1-oxopropan-2-yl)amino)phosphoryl)amino)-2-methylpropanoate;pentyl2-(((R)-((((R)-1-(6-amino-9H-purin-9-yl)propan-2-yl)oxy)methyl)(((R)-1-butoxy-1-oxopropan-2-yl)amino)phosphoryl)amino)-2-methylpropanoate;hexyl2-(((S)-((((R)-1-(6-amino-9H-purin-9-yl)propan-2-yl)oxy)methyl)(((R)-1-isobutoxy-1-oxopropan-2-yl)amino)phosphoryl)amino)-2-methylpropanoate;hexyl2-(((R)-((((R)-1-(6-amino-9H-purin-9-yl)propan-2-yl)oxy)methyl)(((R)-1-isobutoxy-1-oxopropan-2-yl)amino)phosphoryl)amino)-2-methylpropanoate;hexyl2-(((S)-((((R)-1-(6-amino-9H-purin-9-yl)propan-2-yl)oxy)methyl)(((R)-1-butoxy-1-oxopropan-2-yl)amino)phosphoryl)amino)-2-methylpropanoate;hexyl2-(((R)-((((R)-1-(6-amino-9H-purin-9-yl)propan-2-yl)oxy)methyl)(((R)-1-butoxy-1-oxopropan-2-yl)amino)phosphoryl)amino)-2-methylpropanoate;hexyl2-(((S)-((((R)-1-(6-amino-9H-purin-9-yl)propan-2-yl)oxy)methyl)(((R)-1-oxo-1-(pentyloxy)propan-2-yl)amino)phosphoryl)amino)-2-methylpropanoate;hexyl2-(((R)-((((R)-1-(6-amino-9H-purin-9-yl)propan-2-yl)oxy)methyl)(((R)-1-oxo-1-(pentyloxy)propan-2-yl)amino)phosphoryl)amino)-2-methylpropanoate;hexyl2-(((S)-((((R)-1-(6-amino-9H-purin-9-yl)propan-2-yl)oxy)methyl)(((R)-1-(hexyloxy)-1-oxopropan-2-yl)amino)phosphoryl)amino)-2-methylpropanoate;hexyl2-(((R)-((((R)-1-(6-amino-9H-purin-9-yl)propan-2-yl)oxy)methyl)(((R)-1-(hexyloxy)-1-oxopropan-2-yl)amino)phosphoryl)amino)-2-methylpropanoate;hexyl2-(((S)-((((R)-1-(6-amino-9H-purin-9-yl)propan-2-yl)oxy)methyl)(((R)-1-(cyclopentyloxy)-1-oxopropan-2-yl)amino)phosphoryl)amino)-2-methylpropanoate;hexyl2-(((R)-((((R)-1-(6-amino-9H-purin-9-yl)propan-2-yl)oxy)methyl)(((R)-1-(cyclopentyloxy)-1-oxopropan-2-yl)amino)phosphoryl)amino)-2-methylpropanoate;hexyl2-(((S)-((((R)-1-(6-amino-9H-purin-9-yl)propan-2-yl)oxy)methyl)(((R)-1-(cyclohexyloxy)-1-oxopropan-2-yl)amino)phosphoryl)amino)-2-methylpropanoate;hexyl2-(((R)-((((R)-1-(6-amino-9H-purin-9-yl)propan-2-yl)oxy)methyl)(((R)-1-(cyclohexyloxy)-1-oxopropan-2-yl)amino)phosphoryl)amino)-2-methylpropanoate;pentyl2-(((S)-((((R)-1-(6-amino-9H-purin-9-yl)propan-2-yl)oxy)methyl)(((R)-1-oxo-1-(pentyloxy)propan-2-yl)amino)phosphoryl)amino)-2-methylpropanoate;pentyl2-(((R)-((((R)-1-(6-amino-9H-purin-9-yl)propan-2-yl)oxy)methyl)(((R)-1-oxo-1-(pentyloxy)propan-2-yl)amino)phosphoryl)amino)-2-methylpropanoate;pentyl2-(((S)-((((R)-1-(6-amino-9H-purin-9-yl)propan-2-yl)oxy)methyl)(((R)-1-isopropoxy-1-oxopropan-2-yl)amino)phosphoryl)amino)-2-methylpropanoate;pentyl2-(((R)-((((R)-1-(6-amino-9H-purin-9-yl)propan-2-yl)oxy)methyl)(((R)-1-isopropoxy-1-oxopropan-2-yl)amino)phosphoryl)amino)-2-methylpropanoate;isopropyl2-((S)-((((R)-1-(6-amino-9H-purin-9-yl)propan-2-yl)oxy)methyl)(((R)-1-cyclobutoxy-1-oxopropan-2-yl)amino)phosphoryl)amino)-2-methylpropanoate;isopropyl2-(((R)-((((R)-1-(6-amino-9H-purin-9-yl)propan-2-yl)oxy)methyl)(((R)-1-cyclobutoxy-1-oxopropan-2-yl)amino)phosphoryl)amino)-2-methylpropanoate;pentyl2-(((S)-((((R)-1-(6-amino-9H-purin-9-yl)propan-2-yl)oxy)methyl)(((R)-1-cyclobutoxy-1-oxopropan-2-yl)amino)phosphoryl)amino)-2-methylpropanoate;pentyl2-(((R)-((((R)-1-(6-amino-9H-purin-9-yl)propan-2-yl)oxy)methyl)(((R)-1-cyclobutoxy-1-oxopropan-2-yl)amino)phosphoryl)amino)-2-methylpropanoate;isobutyl2-(((S)-((((R)-1-(6-amino-9H-purin-9-yl)propan-2-yl)oxy)methyl)(((R)-1-cyclobutoxy-1-oxopropan-2-yl)amino)phosphoryl)amino)-2-methylpropanoate;isobutyl2-(((R)-((((R)-1-(6-amino-9H-purin-9-yl)propan-2-yl)oxy)methyl)(((R)-1-cyclobutoxy-1-oxopropan-2-yl)amino)phosphoryl)amino)-2-methylpropanoate;hexyl2-(((S)-((((R)-1-(6-amino-9H-purin-9-yl)propan-2-yl)oxy)methyl)(((R)-1-cyclobutoxy-1-oxopropan-2-yl)amino)phosphoryl)amino)-2-methylpropanoate;hexyl2-(((R)-((((R)-1-(6-amino-9H-purin-9-yl)propan-2-yl)oxy)methyl)(((R)-1-cyclobutoxy-1-oxopropan-2-yl)amino)phosphoryl)amino)-2-methylpropanoate;propyl2-(((S)-((((R)-1-(6-amino-9H-purin-9-yl)propan-2-yl)oxy)methyl)(((R)-1-cyclobutoxy-1-oxopropan-2-yl)amino)phosphoryl)amino)-2-methylpropanoate;propyl2-(((R)-((((R)-1-(6-amino-9H-purin-9-yl)propan-2-yl)oxy)methyl)(((R)-1-cyclobutoxy-1-oxopropan-2-yl)amino)phosphoryl)amino)-2-methylpropanoate;butyl2-(((S)-((((R)-1-(6-amino-9H-purin-9-yl)propan-2-yl)oxy)methyl)(((R)-1-cyclobutoxy-1-oxopropan-2-yl)amino)phosphoryl)amino)-2-methylpropanoate;butyl2-(((R)-((((R)-1-(6-amino-9H-purin-9-yl)propan-2-yl)oxy)methyl)(((R)-1-cyclobutoxy-1-oxopropan-2-yl)amino)phosphoryl)amino)-2-methylpropanoate;pentyl2-(((S)-((((R)-1-(6-amino-9H-purin-9-yl)propan-2-yl)oxy)methyl)(((R)-1-(cyclopentyloxy)-1-oxopropan-2-yl)amino)phosphoryl)amino)-2-methylpropanoate;pentyl2-(((R)-((((R)-1-(6-amino-9H-purin-9-yl)propan-2-yl)oxy)methyl)(((R)-1-(cyclopentyloxy)-1-oxopropan-2-yl)amino)phosphoryl)amino)-2-methylpropanoate;pentyl2-(((S)-((((R)-1-(6-amino-9H-purin-9-yl)propan-2-yl)oxy)methyl)(methyl((R)-1-oxo-1-(pentyloxy)propan-2-yl)amino)phosphoryl)amino)-2-methylpropanoate;pentyl2-(((R)-((((R)-1-(6-amino-9H-purin-9-yl)propan-2-yl)oxy)methyl)(methyl((R)-1-oxo-1-(pentyloxy)propan-2-yl)amino)phosphoryl)amino)-2-methylpropanoate;pentyl(((S)-(((R)-1-(6-amino-9H-purin-9-yl)propan-2-yl)oxy)methyl)((2-methyl-1-oxo-1-(pentyloxy)propan-2-yl)amino)phosphoryl)-D-prolinate;pentyl(((R)-(((R)-1-(6-amino-9H-purin-9-yl)propan-2-yl)oxy)methyl)((2-methyl-1-oxo-1-(pentyloxy)propan-2-yl)amino)phosphoryl)-D-prolinate;cyclohexyl2-(((S)-((((R)-1-(6-amino-9H-purin-9-yl)propan-2-yl)oxy)methyl)(((R)-1-(cyclopentyloxy)-1-oxopropan-2-yl)amino)phosphoryl)amino)-2-methylpropanoate;cyclohexyl2-(((R)-((((R)-1-(6-amino-9H-purin-9-yl)propan-2-yl)oxy)methyl)(((R)-1-(cyclopentyloxy)-1-oxopropan-2-yl)amino)phosphoryl)amino)-2-methylpropanoate;cyclobutyl2-(((S)-((((R)-1-(6-amino-9H-purin-9-yl)propan-2-yl)oxy)methyl)(((R)-1-(cyclopentyloxy)-1-oxopropan-2-yl)amino)phosphoryl)amino)-2-methylpropanoate;cyclobutyl2-(((R)-((((R)-1-(6-amino-9H-purin-9-yl)propan-2-yl)oxy)methyl)(((R)-1-(cyclopentyloxy)-1-oxopropan-2-yl)amino)phosphoryl)amino)-2-methylpropanoate;a pharmaceutically acceptable salt thereof.
 4. The compound of claim 2which is

or a pharmaceutically acceptable salt thereof.